Drug Resistance Profiles Research Articles

Circulation of a Unique Klebsiella pneumoniae Clone, ST147 NDM-1/OXA-48, in Two Diverse Hospitals in Calabria (Italy)

Background/Objectives: Carbapenem-resistant Klebsiella pneumoniae has become endemic in Europe, including in Italy, where its prevalence has risen dramatically, primarily due to epidemic clones harboring metallo-enzymes. This study aims to investigate the dissemination of K. pneumoniae strains co-producing OXA-48 and NDM-1 between two hospitals in southern Italy using molecular analyses. Methods: A total of 49 K. pneumoniae strains, predominantly co-producing OXA-48 and NDM-1, were collected between March and December 2023. Antibiotic susceptibility testing was conducted following EUCAST guidelines. Whole-genome sequencing (Illumina MiSeq) and bioinformatics tools (CARD, CLC Genomics Workbench) were used to identify resistance and virulence genes, capsule loci, and phylogenetic relationships. Results: All isolates exhibited multidrug-resistant or extensively drug-resistant profiles, including resistance to ceftazidime/avibactam and meropenem/vaborbactam. Genomic analysis revealed diverse resistance genes such as blaOXA-48, blaNDM-1, blaCTX-M-15, and blaSHV variants. Virulence genes associated with capsules, fimbriae, and siderophores were widespread. Most strains were classified as ST147 by MLST and contained various plasmids known to carry antimicrobial resistance. Phylogenetic analysis confirmed their clonal relatedness, highlighting the intra-hospital dissemination of high-risk clones. Conclusions: High-risk K. pneumoniae clones, particularly ST147, pose significant challenges in healthcare settings due to the extensive antimicrobial resistance driven by plasmid-borne resistance genes, including those that co-produce carbapenemases, like blaNDM-1 and blaOXA-48. Molecular monitoring of these clones is essential for improving targeted infection control strategies, mitigating the spread of multidrug-resistant pathogens, and managing their clinical impact effectively.

Design and Discovery of Preclinical Candidate LYG-409 as a Highly Potent and Selective GSPT1 Molecular Glue Degraders.

Molecular glue degraders induce "undruggable" protein degradation by a proximity-induced effect. Inspired by the clinical success of immunomodulatory drugs, we aimed to design novel molecular glue degraders targeting GSPT1. Here, we report the design of a series of GSPT1 molecular glue degraders. LYG-409, a 2H-chromene derivative, was identified as a potent, selective, and orally bioavailable GSPT1 degrader with excellent antitumor activity in vivo (anti-Acute Myeloid Leukemia MV4-11 xenograft model: TGI = 94.34% at 30 mg/kg; prostate cancer 22Rv1 xenograft model: TGI = 104.49% at 60 mg/kg) and in vitro (KG-1 cells: IC50 = 9.50 ± 0.71 nM, DC50 = 7.87 nM) mediated by the degradation of GSPT1. In conclusion, LYG-409 exhibits potent GSPT1 degradation activity, demonstrating promising therapeutic efficacy and favorable safety profile. However, its potential drug resistance profile needs to be thoroughly evaluated in comparison with existing treatments. We hope LYG-409 can provide a valuable direction for the development of GSPT1 degraders.

Nonclinical and clinical characterization of the absorption, metabolism, and excretion of islatravir.

The development of new and improved antiretroviral therapies that allow for alternative dosing schedules is needed for people living with HIV-1. Islatravir is a deoxyadenosine analog in development for the treatment of HIV-1 that suppresses HIV-1 replication via multiple mechanisms of action, including reverse transcriptase translocation inhibition and delayed chain termination. Islatravir is differentiated from other HIV-1 antiretrovirals by its high potency, long t½, broad tissue distribution, and favorable drug resistance profile. A comprehensive evaluation was performed to provide data on the mass balance, absorption, metabolism, and excretion of islatravir through studies in nonclinical species, and in adults without HIV-1 infection, using radiolabeled islatravir. Islatravir was well absorbed in both nonclinical species and humans following oral administration. The elimination of islatravir occurs primarily by a combination of oxidative deamination to 4'-ethynyl-2-fluoro-2'-deoxyinosine and renal excretion of unchanged islatravir. Islatravir and 4'-ethynyl-2-fluoro-2'-deoxyinosine are the major circulating drug components in all species assessed. Islatravir is readily taken up into cells with efficient phosphorylation to the mono-, di-, and triphosphate forms. The pharmacologically active islatravir triphosphate is the most abundant intracellular phosphorylated species, as shown by the results of ex vivo studies. This characterization of the absorption, metabolism, and elimination of islatravir in humans and nonclinical species supports its further development for the treatment of HIV-1.

HIV Drug Resistance Profile in Clients Experiencing Treatment Failure After the Transition to a Dolutegravir-Based First-Line Antiretroviral Treatment Regimen in Mozambique.

Real-world data on HIV drug resistance (HIVDR) after transitioning to tenofovir disoproxil fumarate/lamivudine/dolutegravir (TLD) are limited. We assessed HIVDR rates and patterns in clients with virological failure (VF) after switching from an NNRTI-based regimen to TLD. A cross-sectional study was conducted in Gaza, Mozambique (August 2021-February 2022), including adults on first-line ART for ≥12 months who transitioned to TLD and had unsuppressed viral load (VL) ≥ 1000 copies/mL six months post-transition. After three adherence counseling sessions, participants with VF underwent genotyping for drug resistance mutations (DRMs) using the Stanford HIVdb Program. Of 717 participants (median age 39.2 years, 70.7% female), 217 (30.2%) had VF, 193 (88.9%) underwent genotyping, with 183 (94.8%) successfully genotyped. Intermediate-high dolutegravir (DTG) resistance was found in 19.6% (36/183). Unsuppressed VL before DTG transition was independently associated with VF (aOR: 2.14). Resistance patterns included 33.3% (12/36; 95% CI: 14.6-46.3) to all three TLD drugs, 55.6% (20/36; 95% CI: 39.3-71.9) to DTG and 3TC, and 11% (4/36; 95% CI: 0.8-21.3) to DTG only. Major drug resistance mutations to DTG included G118R (9.3%), R263K (6.6%), and Q148H/R/K (4.4%). This study highlights the need to consider virologic status before transitioning PLHIV to TLD and suggests that adherence counseling may not prevent resistance in those with unknown or prior VF.

Population Structure Based on Microsatellite Length Polymorphism, Antifungal Susceptibility Profile, and Enzymatic Activity of Candida auris Clinical Isolates in Russia.

Candida auris is an emerging multidrug-resistant fungal pathogen causing nosocomial transmission and invasive infections with high mortality. This study aimed to investigate the genetic relationships, enzymatic activities, and drug-resistance profiles of C. auris isolates to evaluate the population and epidemiological diversity of candidiasis in Russia. A total of 112 clinical isolates of C. auris were analyzed from May 2017 to March 2023 in 18 hospitals across Saint Petersburg, the Leningrad Region, and Moscow. Species identification was confirmed by ITS sequencing, and genotyping was performed using 12 short tandem repeat (STR) markers. Antifungal susceptibility was tested using Sensititre™ YeastOne™ plates, and hydrolytic enzyme production was measured by the plate method. ITS sequencing confirmed that all isolates belonged to a single ITS cluster (clades I and III). Fifteen distinct STR genotypes were identified, with genotype I being dominant (n = 53). The most variable of the analyzed markers turned out to be M3-Ia, which was represented in the Russian population by eight different variants. Fluconazole resistance was found in 111 isolates, 17% were resistant to amphotericin B, and 3.6% to 5-flucytosine. Phospholipase activity was strong in most strains, especially in urine isolates (p = 0.014). Conclusion: The predominance of STR genotype I and its variability at the M3-Ia locus suggest its association with nosocomial outbreaks and transmissibility in Russia.

Drug resistant Mycobacterium tuberculosis strains have altered cell envelope hydrophobicity that influences infection outcomes in human macrophages

In recent decades, drug resistant (DR) strains of Mycobacterium tuberculosis (M.tb), the cause of tuberculosis (TB), have emerged that threaten public health. Although M.tb‘s complex and protective cell envelope has been widely studied, little is known about how levels of peripheral lipids change in relation to drug resistance. In this study, we examined levels of cell envelope lipids [phthiocerol dimycocerosates (PDIMs)], glycolipids [phosphatidyl-myo-inositol mannosides (PIMs)], and PIMs associated lipoglycans [lipomannan (LM); mannose-capped lipoarabinomannan (ManLAM)] of 22 M.tb strains that ranged in drug resistance profile. We show that the PDIMs:PIMs ratio increases as drug resistance increases, and provide evidence of PDIM isomers only present in the DR-M.tb strains studied. Overall, the LM and ManLAM levels did not differ between drug resistance categories, but ManLAM surface exposure increased with drug resistance. Infection of human macrophages revealed that DR-M.tb strains have decreased association compared to drug susceptible (DS) strains, and that the pre-XDR M.tb strain with the largest PDIMs:PIMs ratio had decreased uptake, but increased intracellular growth at early during infection compared to the DS-M.tb strain H37Rv. These findings suggest that PDIMs may play an important role in drug resistance and that an increase in hydrophobic cell envelope lipids may influence M.tb-host interactions.

Antimicrobial resistance of Escherichia coli in cats and their drinking water: drug resistance profiles and antimicrobial-resistant genes

BackgroundAntimicrobial resistance (AMR) is a global health concern that is exacerbated by the transmission of bacteria and genetic material between humans, animals and the environment. This study investigated AMR of Escherichia coli (E. coli) isolated from cats’ feces and their drinking water. The study compared the AMR of fecal and environmental E. coli isolates from pet cats.ResultsA total of 104 samples (52 cat feces and 52 cat drinking water samples) was cultured for E. coli. The study compared the AMR of fecal and environmental E. coli isolates from pet cats. An analysis of carbapenemase and extended-spectrum β-lactamase (ESBL)-producing E. coli genes (blaTEM, blaSHV and blaCTX-M) and phylogroups of E. coli was also performed. E. coli was identified from all fecal (100%) and almost half of drinking water (44.2%) samples. All E. coli isolate was susceptible to amikacin or imipenem. Clindamycin showed the highest resistance rate. β-lactam was the most found with co-resistance profiles, comprising β-lactams with aminoglycosides, quinolones, sulfonamides, macrolides or carbapenems. Very strong positive correlations of bactericidal agents were found among quinolones (r > 0.8, p < 0.01). Within the group of bacteriostatic agents, moderate correlation was observed between azithromycin and sulfa-trimethoprim (r = 0.5253, p < 0.01). Carbapenemase gene was not detected in this study. Extended-spectrum β-lactamase-producing E. coli genes (blaTEM, blaSHV and blaCTX-M) were identified in E. coli isolates, with blaTEM being the most predominant. Furthermore, phylogroup B2 was the dominant segregation among the E. coli, particularly in fecal isolates.ConclusionsThis study identified AMRin E. coli isolated from cats’ feces and their drinking water. The results revealed that the phylogroup B2 was predominant, with blaTEM being the most widespread ESBL gene.

Performance of a broth microdilution assay for routine minimum inhibitory concentration determination of 14 anti-tuberculous drugs against the Mycobacterium tuberculosis complex based on the EUCAST reference protocol.

This comparative study aimed at qualifying a broth microdilution (BMD) assay for phenotypic drug susceptibility testing (pDST) of Mycobacterium tuberculosis complex (MTBC) strains for implementation in a routine DST workflow. The assay was developed based on the EUCAST (European Committee on Antimicrobial Susceptibility Testing) reference protocol for determination of the minimum inhibitory concentration (MIC) of 14 anti-tuberculous drugs (isoniazid [INH], rifampicin [RIF], ethambutol [EMB], amikacin [AMI], moxifloxacin [MFX], levofloxacin [LFX], bedaquiline [BDQ], clofazimine [CFZ], delamanid [DLM], pretomanid [PA], para-aminosalicylic acid [PAS], linezolid [LZD], ethionamide [ETH], and cycloserine [CS]). Forty MTBC strains with various drug resistance profiles were tested to determine the agreement between MIC results and genotypic drug susceptibility testing (gDST) results derived from whole-genome sequencing (WGS). The agreement between the BMD and gDST results was solid for the majority of the drugs (average agreement 98%, range 90%-100%), including key drugs such as INH, RIF, MFX, LFX, BDQ, DLM, and PA. Ten discrepancies were identified (corresponding to 1.8% of the total number of MIC determinations) and most (8/10) were characterized by MICs equal or close to the potential critical concentration (pCC) applied in the BMD assay. Importantly, the assay can be adjusted to new drug recommendations and concentrations, tailored to local needs. We conclude that the BMD assay provides reliable results, and its implementation in our MTBC routine workflow will produce valuable data that improve our understanding and management of MTBC drug resistance.

Drug-resistant pulmonary tuberculosis with surgical treatment: clinical forms, histological and macroscopic aspects

OBJECTIVE. To clarify the clinical forms of drug-resistant pulmonary tuberculosis with surgical treatment and their relationship with the morphological activity of specific inflammation and the detection of Mycobacterium tuberculosis (MBT) in the surgical material. MATERIALS AND METHODS. The study had a cohort retrospective character. The two groups of patients were studied – with multidrug-resistant tuberculosis (n=27) and pre-extensively drug-resistant tuberculosis (n=25). All patients after completion of a standardized 6-month or individualized chemotherapy was performed surgery. The results of morphological examination namely clarification of pulmonary tuberculosis clinical forms and the morphological degree of activity of a specific inflammatory process were analyzed. A comparative analysis of MBT detection by the cultural method in the surgical material depending on the forms of pulmonary tuberculosis and its degree of activity was performed. RESULTS AND DISCUSSION. Regardless of the drug-resistant profile, different forms of pulmonary tuberculosis were presented in both study groups. In general, cases of multiple tuberculomas (24; 46.2 %) were quantitative prevailed with predominance in group 1. In second place in frequency was diagnosed fibrocaseous tuberculosis (10; 19.2 %) at that cases of pre-extensively drug-resistant tuberculosis (7; 28.0 %) more than twice prevailed. A diagnosis of residual post-tuberculosis changes in the lungs (7; 13.5 %) was at third place, which also prevailed in group 2 (6; 24.0 %). According of morphological examination, it was found that only a quarter of patients at the time of surgery achieved remission of the disease (12; 23.1 %) with a predominance of group 1 (8; 32.0 %). The progression of specific inflammation persisted (18; 34.6 %) in a third of observations in both groups. During surgery operating material was collected for cultural research, which was conducted in 41 cases (78.8 %). In most cases (30; 57.7 %), MBT was not detected, the largest proportion occurred on lung tuberculoma (17; 32.7 %). In almost quarter of all cases, the MBT was detected in surgical material (12; 23.1 %). The share of MBT detection was much higher in fibrocaseous tuberculosis (4; 40.0 %), compared to cases of solitary tuberculomas (4; 16.7 %). The MBT was also detected in residual post-tuberculosis changes – only in the group of pre-extensively drug-resistant tuberculosis (3; 12.0 %). It is established that the morphological activity of the inflammatory process does not clearly correlate with cases of cultural detection of MBT. CONCLUSIONS. The use of modern chemotherapeutic regimens for the treatment of tuberculosis with multidrug resistance and pre-extensively drug-resistant tuberculosis during, in the main course of therapy does not allow to achieve complete abacylation in such patients, which causes additional surgical treatment. In cases of pre-extensively drug-resistant tuberculosis, the development of more severe forms of lung tuberculosis and the lower preoperative effectiveness of specific anti-tuberculosis therapy was observed.

Genotypes and drug resistance pattern of Mycobacterium tuberculosis complex among clinically diagnosed pulmonary tuberculosis patients.

Clinically diagnosed pulmonary tuberculosis (TB) (CDPTB) patients account for a huge proportion of TB. However, little is known about the genetic diversity and drug resistance profile of Mycobacterium tuberculosis Complex (MTBC) strains in this group of patients. Unmatched case-control study was conducted among 313 PTB patients to compare the genetic diversity of MTBC and their drug resistance profiles among CDPTB (n = 173) and bacteriologically confirmed pulmonary TB (BCPTB) (n = 140) patients. Lowenstein-Jensen (LJ) culture, geneXpert and acid fast staining were performed on sputum specimen collected from both CDPTB and BCPTB patients. Spoligotyping, whole genome sequencing (WGS) and phenotypic drug resistance testing (DST) were done for a subset of LJ grown MTBC isolates. Data was analyzed by STATA version 17 software and a p-value <0.05 were considered statistically significant. The proportion of lineage 3 was larger among CDPTB patients (31%, 13/42) compared to BCPTB patients (15%, 11/74) (p-value <0.05). A higher proportion of MTBC isolates from CDPTB 16.6% (3/18) were phenotypically resistant to one or more anti-TB drugs than BCPTB 12% (4/33) (p-value >0.05). A single lineage 3 strain resistant to all the primary anti-TB drugs was detected in one CDPTB by both DST methods. The observed differences in the genotypes of MTBC isolates between CDPTB and BCPTB patients may be attributed to challenges in the identification of CDPTB that requires further investigation on sequenced genome of the MTBC strains for better understanding and recommendation based on the current finding. There was also primary drug resistant TB among culture positive CDPTB patients which would be otherwise missed by current national protocols.

Comparative Genomic Analysis of an Apiotrichum cacaoliposimilis Strain Isolated from a Patient with Urinary Tract Infection.

Opportunistic infections caused by fungi, particularly those occurring in immunocompromised patients, are considered challenging worldwide. Therefore, a comprehensive understanding of pathogenic fungi is necessary. The present study reports the isolation of a strain of Apiotrichum cacaoliposimilis, which is difficult to detect using conventional clinical assays, from the sterile urine samples of a patient with a urinary tract infection. Sanger sequencing of the internal transcribed spacer regions confirmed the genus of the microbe, while whole-genome sequencing yielded the initial genome assembly of A. cacaoliposimilis. A total of 7,161 predicted proteincoding genes were mapped using multiple databases, including Gene Ontology, Kyoto Encyclopedia of Genes and Genomes, non-redundant protein database, Pathogen-Host Interactions Database, and Comprehensive Antibiotic Resistance Database. The phenotypic data, biochemical reactions, and antimicrobial susceptibility analyses were conducted to reveal the metabolic properties, virulence, and drug resistance profile of the isolated A. cacaoliposimilis. The rank-sum test revealed the differences in the intergeneric distribution of the highly virulent genes UgeB and Pem2. In addition, other genes exhibited significant overlap in terms of virulence factors with the clinical isolate Apiotrichum mycotoxinivorans GMU1709. Fortunately, similar to most fungi belonging to the Apiotrichum genus, the isolate investigated in the present study was also sensitive to the drug voriconazole (MIC = 0.06 μg/ml). In summary, the phylogenetic placement, potential pathogenic genes, drug sensitivity patterns, and morphological characteristics of the isolated A. cacaoliposimilis were determined precisely in the present study.

Combination of gallium citrate and levofloxacin induces a distinct metabolome profile and enhances growth inhibition of multidrug-resistant Mycobacterium tuberculosis compared to linezolid.

Tuberculosis (TB) treatment typically involves a tailored combination of four antibiotics based on the drug resistance profile of the infecting strain. The increasing drug resistance of Mycobacterium tuberculosis (Mtb) requires the development of novel antibiotics to ensure effective treatment regimens. Gallium (Ga) is being explored as a repurposed drug against TB due to its ability to inhibit Mtb growth and disrupt iron metabolism. Given the potential interactions between Ga and established antibiotics, we investigated how a combination of Ga with levofloxacin (Lfx) or linezolid (Lzd) affects the growth and metabolome of a multidrug-resistant (MDR) Mtb clinical strain. Mtb was cultured using a BACTEC 960 system with concentrations of Ga ranging from 125 to 1,000 μM and with 250 to 500 μM of Ga combined with 0.125 mg/L of Lfx or Lzd. For metabolome analysis, the antibacterials were used at concentrations that inhibited the growth of bacteria without causing cell death. Metabolites were extracted from Mtb cells and analyzed using chromatography-mass spectrometry. The MDR Mtb strain exhibited a dose-dependent response to Ga. Notably, the enhancement in growth inhibition was statistically significant for the Ga/Lfx combination compared to Ga alone, while no such significance was observed for Ga/Lzd. Moreover, exposure to Ga/Lfx or Ga/Lzd resulted in distinct metabolite profiles. Ga treatment increased the level of aconitate, fumarate, and glucose in the cells, suggesting the inhibition of iron-dependent aconitase and fumarate hydratase, as well as disruption of the pentose phosphate pathway. The levels of glucose, succinic acid, citric acid, and hexadecanoic acid followed a similar pattern in cells exposed to Ga and Ga/Lfx at 500 μM Ga but exhibited different trends at 250 μM Ga. In the presence of Lfx, the Mtb metabolome changes induced by Ga are more pronounced compared to those observed with Lzd. Lfx affects nucleic acids and transcription, which may enhance Ga-dependent growth inhibition by preventing the metabolic redirection that bacteria typically use to bypass iron-dependent enzymes.

Etiología y resistencia a los antibióticos en la infección urinaria pediátrica. Estudio multicéntrico desde Atención Primaria

Introduction: empirical treatment of urinary tract infection (UTI) requires knowledge of antibiotic resistance in the most prevalent uropathogens. The aim of the study was to analyse the pathogens involved in community-acquired paediatric UTIs, their drug resistance profile and the association between resistance and the studied variables. Patients and methods: nationwide, multicentre, prospective, longitudinal and descriptive study carried out in primary care paediatrics clinics. We included all UTI episodes identified in patients aged 0-15 years in the caseloads of 187 collaborating paediatricians (187 058 patients), regardless of the setting where they were diagnosed or treated, between 1/10/2019 and 31/12/2020. Results: there were 588 identified UTI episodes. Escherichia coli was the most frequently isolated uropathogen (79.67%), followed by Proteus spp., Klebsiella spp. and Enterococcus spp. Extended-spectrum beta-lactamase (ESBL)-producing Enterobacteriaceae were isolated in 1.52% of episodes. The lowest prevalences of resistance to oral antibiotics corresponded to third-generation cephalosporins, cefuroxime and fosfomycin (3.88%, 4.81% and 6.30%, respectively). The proportions of resistance to first-generation cephalosporins, gentamicin and amikacin were 8.69%, 7.55% and 3.23% respectively. The prevalence of resistance was 23.85% for amoxicillin-clavulanic acid (34.51% in males) and 23.40% for cotrimoxazole. Conclusions: E. coli was the most frequently isolated uropathogen. Amoxicillin-clavulanic acid and cotrimoxazole should not be used empirically due to the high prevalence of resistance. Second and third generation cephalosporins and fosfomycin could be adequate empirical therapy depending on age and the type of infection. Local susceptibility to first-generation cephalosporins should be tested.

An Oxford Nanopore Technology-Based Hepatitis B Virus Sequencing Protocol Suitable for Genomic Surveillance Within Clinical Diagnostic Settings.

Chronic Hepatitis B Virus (HBV) infection remains a significant public health concern, particularly in Africa, where the burden is substantial. HBV is an enveloped virus, classified into ten phylogenetically distinct genotypes (A-J). Tests to determine HBV genotypes are based on full-genome sequencing or reverse hybridization. In practice, both approaches have limitations. Whereas diagnostic sequencing, generally using the Sanger approach, tends to focus only on the S-gene and yields little or no information on intra-patient HBV genetic diversity, reverse hybridization detects only known genotype-specific mutations. To resolve these limitations, we developed an Oxford Nanopore Technology (ONT)-based HBV diagnostic sequencing protocol suitable for clinical virology that yields both complete genome sequences and extensive intra-patient HBV diversity data. Specifically, the protocol involves tiling-based PCR amplification of HBV sequences, library preparation using the ONT Rapid Barcoding Kit (Oxford nanopore Technologies, Oxford, OX4 4DQ, UK), ONT GridION sequencing, genotyping using genome detective software v1.132/1.133, a recombination analysis using jpHMM (26 October 2011 version) and RDP5.61 software, and drug resistance profiling using Geno2pheno v2.0 software. We prove the utility of our protocol by efficiently generating and characterizing high-quality near full-length HBV genomes from 148 residual diagnostic samples from HBV-infected patients in the Western Cape province of South Africa, providing valuable insights into the genetic diversity and epidemiology of HBV in this region of the world.

Drug resistance profiling, antimicrobial susceptibility and demographic characteristics of children with acute bacterial meningitis in a Southeastern tertiary health facility

Background: Over the years, varying patterns of bacterial susceptibility and multidrug resistance (MDR) rates have been reported in different settings. Detailed evaluation of the drug profile of the bacterial pathogens implicated in children with invasive bacterial infection helps to reduce the heightened risk of adverse events that could follow poorly managed or complicated cases in resource limited environment like ours. This study therefore, aimed to highlight the antibiotic susceptibility and MDR rate, and establish the relationship if any, between demographic characteristics and positive CSF- bacterial isolates of post neonatal children with suspected acute bacterial meningitis (ABM). The findings would guide practitioners on the empirical antimicrobials to consider in the event of clinical suspicion of ABM pending the availability of CSF isolates' antibiogram. Methods: A prospective review of 100 children with clinical suspicion of ABM from January 2016- December 2020. Descriptive statistics, chi square and regression analysis were used to establish MDR rates, Isolates' susceptibility pattern and the relationship between demographic variables and positive isolates respectively. P&lt;0.05 was accepted as significant. Results: Fluroquinolones, cephalosporins, imipenem and aminoglycosides were susceptible anti-microgram in children with ABM. Sixty-four (85.5%) of the isolates showed MDR pattern, and young children (infants and toddlers) were significantly associated with positive CSF bacterial isolates. Conclusions: ABM should be treated with combination of CNS penetrating empirical antibiotics due to rising rate of MDR pathogens. Young children with febrile illnesses should be thoroughly evaluated for possibility of CNS infection.

Improving Predictive Efficacy for Drug Resistance in Novel HIV-1 Protease Inhibitors through Transfer Learning Mechanisms.

The human immunodeficiency virus presents a significant global health challenge due to its rapid mutation and the development of resistance mechanisms against antiretroviral drugs. Recent studies demonstrate the impressive performance of machine learning (ML) and deep learning (DL) models in predicting the drug resistance profile of specific FDA-approved inhibitors. However, generalizing ML and DL models to learn not only from isolates but also from inhibitor representations remains challenging for HIV-1 infection. We propose a novel drug-isolate-fold change (DIF) model framework that aims to predict drug resistance score directly from the protein sequence and inhibitor representation. Various ML and DL models, inhibitor representations, and protein representations were analyzed through realistic validation mechanisms. To enhance the molecular learning capacity of DIF models, we employ a transfer learning approach by pretraining a graph neural network (GNN) model for activity prediction on a data set of 4855 HIV-1 protease inhibitors (PIs). By performing various realistic validation strategies on internal and external genotype-phenotype data sets, we statistically show that the learned representations of inhibitors improve the predictive ability of DIF-based ML and DL models. We achieved an accuracy of 0.802, AUROC of 0.874, and r of 0.727 for the unseen external PIs. By comparing the DIF-based models with a null model consisting of isolate-fold change (IF) architecture, it is observed that the DIF models significantly benefit from molecular representations. Combined results from various testing strategies and statistical tests confirm the effectiveness of DIF models in testing novel PIs for drug resistance in the presence of an isolate.

Analysis of the changes of bacterial spectrum and drug resistance in sputum culture of ICU children in a hospital of pediatric in Jiangsu Province from 2017 to 2022

Objective: To investigate the changes of the distribution and drug resistance profile of bacteria from ICU children with lower respiratory tract infection (LRTI) in Suzhou City, Jiangsu Province from 2017 to 2022. Methods: From January 2017 to December 2022, a cross-sectional observational study on the bacterial spectrum analysis among intensive care unit (ICU) children with LRTI was conducted in Children's Hospital of Soochow University. The bacteria was cultivated by culture methods from sputum samples, and identified by MALDI-TOF mass spectrometry. Drug sensitivity tests were performed by the VITEK2 Compact fully automated analysis system and the paper slide method. The χ2 test or Fisher's exact probability was used to analyze the changes of the distribution of sputum culture-positive bacteria and drug resistance in ICU children. Results: The overall detection rate of sputum culture was 42.06% (1 182/2 810). Staphylococcus aureus (25.63%,303/1 182), Acinetobacter baumannii (13.62%,161/1 182) and Haemaphilus influenzae (13.28%,157/1 182) were the top three. Proportions of Acinetobacter baumannii (17.90% vs. 11.02%,χ²=11.17, P=0.001), especially carbapenem-resistant Acinetobacter baumannii (43.70% vs. 23.50%, χ²=15.21, P<0.001) increased significantly from 2020 to 2022. However, the proportions of Haemophilus influenzae (8.50% vs. 16.19%, χ²=14.27, P<0.001), Streptococcus pneumoniae (8.50% vs. 15.92%, χ²=13.42, P<0.001) and extended-spectrum-lactamase producing Escherichia coli (8.89% vs. 18.00%, χ²=5.45, P=0.025) decreased. Drug resistant results showed that Acinetobacter baumannii was obviously more resistant to imipenem (χ²=4.43, P=0.035) and levofloxacin (χ²=12.53, P<0.001), while more sensitive to minocycline (χ²=8.34, P=0.004). Escherichia coli showed a significant increase in resistance to piperacillin tazobactam (χ²=8.29, P=0.008) and cefoperazone sulbactam (χ²=5.07, P=0.024) from 2020 to 2022; Klebsiella pneumoniae consistently maintained a resistance rate of more than 60% to first and second-generation cephalosporins, and remain susceptible to quinolones and carbapenems. Staphylococcus aureus remained highly susceptible to levofloxacin (drug resistance rate: 2.31%,7/303) and sulfamethoxazole/trimethoprim (drug resistance rate: 4.95%,15/303) from 2020 to 2022. Conclusion: Higher detection and resistance rates of Acinetobacter baumannii from sputum culture in ICU children from 2020 to 2022 were explored. Resistance of Escherichia coli to β-lactamase inhibitor combinations was more serious. Regular monitoring the changes of the etiology of respiratory tract infections in ICU Children is particularly important for the prevention and treatment of multidrug-resistant bacterial infections.

Cutaneous Atypical Mycobacterial Infections: A Brief Review.

Nontuberculous mycobacterial (NTM) infections are increasingly recognized, particularly in tropical regions and are often found in immunocompetent individuals. These infections are emerging as significant health concerns, especially pulmonary NTM, which is reported more frequently and is known to be associated with hospital environments. While pulmonary NTM infections are on the rise, partly due to drug resistance and possible patient-to-patient transmission, there is no current evidence indicating an increase in cutaneous NTM infections. The clinical manifestations of NTM infections, except for well-known entities like Buruli ulcer and fish tank granuloma, are diverse and nonspecific, often mimicking other chronic infections. History of minor trauma at the site of infection can be misleading and may complicate the diagnosis of cutaneous NTM. Surgical-site and port-site NTM infections typically present with erythema, edema, and abscesses and are commonly caused by rapidly growing mycobacteria like M. fortuitum and M. chelonae. These infections may not respond to standard antibiotics, suggesting the need for NTM-specific treatment. Diagnostically, histopathology may not be conclusive, and standard staining techniques often lack sensitivity. Molecular methods offer better speciation and drug resistance profiling for pulmonary NTM but are expensive and not widely available for cutaneous forms. The high cost and limited availability of diagnostic tools necessitate an empirical treatment approach, which is also recommended by the INDEX-Tb guidelines for extrapulmonary tuberculosis. Empirical treatment regimens for NTM, such as combinations of clarithromycin, doxycycline, and cotrimoxazole or fluoroquinolones, have shown promise, but there is a lack of rigorous studies to establish standardized treatments. Monitoring for adverse effects and continued evaluation of the causative organism is essential during empirical treatment, allowing for adjustment if the initial regimen fails.

Pyrazinamide Resistance: A Major Cause of Switching Shorter to Longer Bedaquiline-based Regimens in Multidrug-resistant Tuberculosis Patients.

All-oral regimens, including bedaquiline, are now standard in shorter treatment regimens (STRs) for multidrug-resistant tuberculosis (MDR-TB). Resistance or intolerance to drugs in STR often necessitates a switch to longer treatment regimens (LTRs). This study aims to identify the factors associated with this transition in MDR-TB patients. We conducted a retrospective analysis of medical records from MDR-TB patients treated with STR at Haji Hospital, Surabaya, between January 2022 and January 2023. Data on drug-resistance profiles, determined by drug-susceptibility testing (DST), and line probe assay, as well as adverse effects, were collected. Among 20 eligible patients, 8 (40.0%) switched from STR to LTR within the first 4 months. Resistance was observed in 62.5% of these patients for pyrazinamide, 25.0% for high-dose isoniazid, and 12.5% for levofloxacin. The overall prevalence of pyrazinamide resistance was 25.0%. A history of prior antitubercular treatment was significantly associated with pyrazinamide resistance (P = 0.015; RR - 16.000; confidence interval 95% 1.274-200.917). Pyrazinamide resistance is a major factor for switching from STR to LTR in MDR-TB patients, particularly among those with previous TB treatment. Rapid DST for pyrazinamide is essential for the early identification of resistance and timely adjustments to treatment regimens.

Evaluating culture-free targeted next-generation sequencing for diagnosing drug-resistant tuberculosis: a multicentre clinical study of two end-to-end commercial workflows.

Drug-resistant tuberculosis remains a major obstacle in ending the global tuberculosis epidemic. Deployment of molecular tools for comprehensive drug resistance profiling is imperative for successful detection and characterisation of tuberculosis drug resistance. We aimed to assess the diagnostic accuracy of a new class of molecular diagnostics for drug-resistant tuberculosis. We conducted a prospective, cross-sectional, multicentre clinical evaluation of the performance of two targeted next-generation sequencing (tNGS) assays for drug-resistant tuberculosis at reference laboratories in three countries (Georgia, India, and South Africa) to assess diagnostic accuracy and index test failure rates. Eligible participants were aged 18 years or older, with molecularly confirmed pulmonary tuberculosis, and at risk for rifampicin-resistant tuberculosis. Sensitivity and specificity for both tNGS index tests (GenoScreen Deeplex Myc-TB and Oxford Nanopore Technologies [ONT] Tuberculosis Drug Resistance Test) were calculated for rifampicin, isoniazid, fluoroquinolones (moxifloxacin, levofloxacin), second line-injectables (amikacin, kanamycin, capreomycin), pyrazinamide, bedaquiline, linezolid, clofazimine, ethambutol, and streptomycin against a composite reference standard of phenotypic drug susceptibility testing and whole-genome sequencing. Between April 1, 2021, and June 30, 2022, 832 individuals were invited to participate in the study, of whom 720 were included in the final analysis (212, 376, and 132 participants in Georgia, India, and South Africa, respectively). Of 720 clinical sediment samples evaluated, 658 (91%) and 684 (95%) produced complete or partial results on the GenoScreen and ONT tNGS workflows, respectively, with 593 (96%) and 603 (98%) of 616 smear-positive samples producing tNGS sequence data. Both workflows had sensitivities and specificities of more than 95% for rifampicin and isoniazid, and high accuracy for fluoroquinolones (sensitivity approximately ≥94%) and second line-injectables (sensitivity 80%) compared with the composite reference standard. Importantly, these assays also detected mutations associated with resistance to critical new and repurposed drugs (bedaquiline, linezolid) not currently detectable by any other WHO-recommended rapid diagnostics on the market. We note that the current format of assays have low sensitivity (≤50%) for linezolid and more work on mutations associated with drug resistance is needed. This multicentre evaluation demonstrates that culture-free tNGS can provide accurate sequencing results for detection and characterisation of drug resistance from Mycobacterium tuberculosis clinical sediment samples for timely, comprehensive profiling of drug-resistant tuberculosis. Unitaid.