Beta-lactam Antibiotics Research Articles

Overexpression of lasB Gene in Klebsiella pneumoniae and its Effect on Biofilm Formation and Antibiotic Resistance

Background: Klebsiella pneumoniae is the second most frequent pathogenic bacterium in the Enterobacteriaceae family, after E. coli. It is also regarded as a major pathogen responsible for healthcare-associated infections around the world. Objective: To look at how overexpressing the elastase gene (lasB) in K. pneumoniae affects biofilm development and antibiotic resistance. Methods: 25 clinical isolates of K. pneumoniae were received from Baghdad's Chemistry Analysis Center (CAC) and re-identified using the Vitek-2 method. The microtiter plate was used to measure biofilm production with ELISA. The disc diffusion method was used in antibiotic sensitivity tests in accordance with the CLSI 2022 criteria. The PlasB plasmid was transformed into K. pneumoniae via electroporation. Results: Out of 25 isolates, 11 (44%), 11 (44%), and 3 (12%) produced strong, moderate, and weak biofilms, respectively. One strong biofilm producer (KA1) was chosen for further investigation. The lasB plasmid was successfully transformed into KA1, yielding the KA1 (plasB) isolate. KA1 (plasB) formed considerably fewer biofilms than KA1, and it was more susceptible to tetracycline, doxycycline, and amoxicillin-clavulanic acid than KA1. Furthermore, KA1 (plasB) has shown a significant decrease in ampicillin resistance and an increase in ciprofloxacin sensitivity, but no variations in susceptibility to levofloxacin, cefotaxime, piperacillin-tazobactam, amikacin, or erythromycin when compared to KA1. Conclusions: Overexpression of the elastase gene (plasB) has a major impact on biofilm development and antibiotic resistance in K. pneumoniae.

Prevalence of Extended-Spectrum β-Lactamases (ESBLs) Producing Aeromonas spp. Isolated from Lamellidens marginalis (Lamark, 1819) of Sewage-Fed Wetland: A Phenotypic and Genotypic Approach.

The global rise of zoonotic bacteria resistant to multiple antimicrobial classes and the growing occurrence of infections caused by Aeromonas spp. resistant to β-lactam antibiotics pose a severe threat to animal and human health. However, the contribution of natural environments, particularly aquatic ecosystems, as ideal settings for the development and spread of antimicrobial resistance (AMR) is a key concern. Investigating the phenotypic antibiotic resistance and detection of β-lactamase producing Aeromonas spp. in Lamellidens marginalis, which inhabit all freshwater ecosystems of the Indian subcontinent, is essential for implications in monitoring food safety and drug resistance. In the present investigation, 92 isolates of Aeromonas spp. were recovered from 105 bivalves and screened for their antimicrobial resistance patterns. In vitro antibiotic resistance profiling showed a higher Multiple Antibiotic Resistance (MAR) index of 0.8 with the highest resistance against ampicillin/sulbactam (82%), while 58, 44, 39 and 38% of the isolates were resistant to cephalothin, erythromycin, cefoxitin and imipenem, respectively. PCR results revealed that these isolates carried the blaTEM gene (94%), which was followed by the blaCTX-M gene (51%) and the blaSHV gene (45%). A combination of blaSHV, blaCTX-M, and blaTEM genes was found in 17% of the isolates, indicating the presence of all three resistance genes. This is the first investigation which highlights the importance of multidrug-resistant Aeromonas spp. in L. marginalis. The identification of extended-spectrum-β-lactamases (ESBLs) genes demand the necessity of continuous surveillance and systematic monitoring, considering its potential health risks for both animals and human beings.

Genomic context as well as sequence of both psr and penicillin-binding protein 5 contributes to β-lactam resistance in Enterococcus faecium.

The findings of this study shed light on ampicillin resistance in Enterococcus faecium clade A strains. They underscore the significance of alterations in the amino acid sequence of penicillin-binding protein 5 (PBP5) and the pivotal role of the psr region in PBP5 expression and ampicillin resistance. Notably, the presence of a full-length psrB leads to reduced PBP5 expression and lower minimum inhibitory concentrations (MICs) of ampicillin compared to the presence of a shorter psrA, regardless of the pbp5 allele involved. Additionally, clade B E. faecium strains exhibit lower AMP MICs when both psr alleles from clades A and B are present, although it is important to consider other distinctions between clade A and B strains that may contribute to this effect. It is intriguing to note that the divergence between clade A and clade B E. faecium and the subsequent evolution of heightened AMP MICs in hospital-associated strains appear to coincide with changes in Pbp5 and psr. These changes in psr may have resulted in an inactive Psr, facilitating increased PBP5 expression and greater ampicillin resistance. These results raise the possibility that a mimicker of PsrB, if one could be designed, might be able to lower MICs of ampicillin-resistant E. faecium, thus potentially resorting ampicillin to our therapeutic armamentarium for this species.

Pseudomonas aeruginosa from a Tertiary Teaching Hospital: A Study of AmpC and Biofilm-producing Clinical Isolates

Abstract Introduction: Opportunistic Pseudomonas aeruginosa is one of the most prevalent bacteria with a broad spectrum of human-associated infections. The World Health Organization (WHO) claims that Pseudomonas aeruginosa infection is a typical healthcare-associated disease of critical or high concern. It is well known for being inherently resistant to many antibiotics and has the ability to produce biofilm. In the present study, we aimed to investigate P. aeruginosa with reference to biofilm and AmpC β-lactamases producing isolates in a tertiary care hospital. Materials and Methods: An observational cross-sectional study was performed at the Department of Microbiology, KIMS, KVV, Karad, and Krishna Hospital & Medical Research Center, Karad. (KH & MRC). A total of 180 isolates were subjected to this test to find out which produced the AmpC β -lactamase enzyme by the cefoxitin-cloxacillin double-disc synergy test method and biofilm production by tissue culture plate assay method. Results: Our study found a total of 97 isolates (53.89%) of AmpC production and biofilm production accounted for 151 (83.89%). Conclusion: AmpC- beta-lactamases are responsible for increasing the resistance of P. aeruginosa to several beta-lactam antibiotics and this study also showed that the clinical isolates of P. aeruginosa had a higher propensity to form biofilm and that there was a direct association between biofilm formation and antibiotic resistance.

Beta-Lactam Allergy De-labeling in a Pediatric Hospital.

To assess the ability to de-label pediatric patients of their beta-lactam allergy by using a newly implemented institutional protocol and to identify potential barriers to the de-labeling process. All patients with reported allergies to prespecified beta-lactam antibiotics were eligible for a -beta-lactam allergy interview. Following the interview, patients were grouped into 4 risk categories-no risk, low risk, moderate risk, and high risk-and assessed for intervention eligibility. Potential interventions included de-labeling based on the interview alone or proceeding to an oral amoxicillin challenge with or without penicillin allergy skin testing. Of the 62 patients eligible for beta-lactam allergy interviews, 40% (n = 25) were de-labeled. Among de-labeled patients, 60% (n = 15) were de-labeled on the basis of the interview alone. Additionally, no failures were documented in patients who underwent an oral amoxicillin challenge or penicillin skin testing. Barriers to performing oral amoxicillin challenges or penicillin skin testing included concomitant systemic steroid or antihistamine use, refusal of intervention, and insufficient resources to perform penicillin skin testing. There was a high frequency of patients de-labeled of their beta-lactam allergies in this study. Increased education to patients, parents, and providers on the de-labeling process, as well as increased personnel available to coordinate and perform de-labeling interventions, may result in more beta-lactam allergy de-labeling.

Effects of novel GLT-1 modulator, MC-100093, on neuroinflammatory and neurotrophic biomarkers in mesocorticolimbic brain regions of male alcohol preferring rats exposed chronically to ethanol

Chronic ethanol consumption can lead to increased extracellular glutamate concentrations in key reward brain regions, such as medial prefrontal cortex (mPFC) and nucleus accumbens (NAc), and consequently leading to oxidative stress and neuroinflammation. Previous studies from our lab tested β-lactam antibiotics and novel beta-lactam non-antibiotic, MC-100093, and showed these β-lactam upregulated the major astrocytic glutamate transporter, GLT-1, and consequently reduced ethanol intake and normalized glutamate homeostasis. This present study tested the effects of novel synthetic β-lactam non-antibiotic drug, MC-100093, in chronic ethanol intake and neuroinflammatory and trophic factors in subregions of the NAc (NAc core and shell) and mPFC (Prelimbic, PL; and Infralimbic, IL) of male P rats. MC-100093 treatment reduced ethanol intake after 5-week drinking regimen. Importantly, MC-100093 attenuated ethanol-induced downregulation of brain derived neurotrophic factor (BDNF) expression in these brain regions. In addition, MC-100093 attenuated ethanol-induced upregulation of pro-inflammatory cytokines such as TNF-a and HMGB1 in all these brain regions. Furthermore, MC-100093 treatment attenuated ethanol-induced increase in RAGE in these brain regions. MC-100093 prevented neuroinflammation caused by ethanol intake as well as increased neurotrophic factor in mesocorticolimbic brain regions. MC-100093 treatment reduced ethanol intake and this behavioral effect was associated with attenuation of reduced trophic factors and increased pro-inflammatory factors. MC-100093 is considered a small molecule that may have potential therapeutic effects for the treatment of the effects of chronic exposure to ethanol.

Antibiotic-Resistance Profiles and Genetic Diversity of Shigella Isolates in China: Implications for Control Strategies.

The urgent need for comprehensive and systematic analyses of Shigella as the key pathogen led us to meticulously explore the epidemiology and molecular attributes of Shigella isolates. Accordingly, we procured 24 isolates (10 from Xinjiang and 14 from Wuhan, China) and performed serotype identification and antimicrobial susceptibility testing. Resistance gene detection and homology analysis by polymerase chain reaction and pulsed-field gel electrophoresis (PFGE), respectively, were performed for genetic diversity analysis. All isolates were identified as Shigella flexneri, with 70% (35.4-91.9%) and 30% (8.1-64.6%) of the Xinjiang isolates and 85.7% (56.2-97.5%) and 14.3% (2/14, 2.5-43.9%) of the Wuhan isolates belonging to serotype 2a and serotype 2b, respectively. All isolates displayed resistance to at least two antibiotics and complete resistance to ampicillin. Multidrug resistance (MDR) was recorded in 70.8% (48.8-86.6%) of isolates, with Xinjiang isolates exhibiting relatively higher resistance to ampicillin-sulbactam, piperacillin, ceftriaxone, and aztreonam. Conversely, Wuhan isolates displayed higher MDR and resistance to tetracycline, ciprofloxacin, levofloxacin, and cefepime relative to Xinjiang isolates. Molecular scrutiny of antibiotic-resistance determinants revealed that blaTEM was the main mechanism of ampicillin resistance, blaCTX-M was the main gene for resistance to third- and fourth-generation cephalosporins, and tetB was the predominant gene associated with tetracycline resistance. Four Xinjiang and seven Wuhan isolates shared T1-clone types (>85%), and two Xinjiang and one Wuhan isolates were derived from the T6 clone with a high similarity of 87%. Six PFGE patterns (T1, T2, T5, T6-3, T8, and T10) of S. flexneri were associated with MDR. Thus, there is a critical need for robust surveillance and control strategies in managing Shigella infections, along with the development of targeted interventions and antimicrobial stewardship programs tailored to the distinct characteristics of Shigella isolates in different regions of China.

Contribution of qPCR to the diagnosis of cervico-vaginal infections at the Hôpital Principal de Dakar, Senegal

To determine the etiology of cervico-vaginal infections by cytobacteriology and the efficacy of qPCR for the diagnosis of sensitive strains such as Streptococcus agalactiae, Borrelia crocidurae, Chlamydia trachomatis, Neisseria gonorrhoeae and Treponema pallidum. This prospective cross-sectional study was performed between January and September 2021 in 346 women who were examined for cervico-vaginal infection at the Hôpital Principal de Dakar (HPD). Cytobacteriological (direct examination, agar culture) and molecular analyses were performed. Vaginal flora imbalances predominated, with a rate of 72.3%. The proportion of type IV vaginal flora was 46.5%. Of the 199 germs isolated, Candida albicans (25.1%), Ureaplasma urealyticum (17.6%), S. agalactiae (7.8%), Gardnerella vaginalis (6.6%) and nonalbicans Candida (5.5%) were the main pathogens responsible for cervico-vaginal infections in patients. Among women tested for mycoplasma, U. urealyticum was identified in 43.3% of patients. Among those tested for C. trachomatis, the proportion of infected women was low (4%). The prevalence of C. albicans was higher in pregnant women (38.3%) than in nonpregnant women (19.2%). S. agalactiae strains showed high resistance to certain beta-lactam antibiotics (pristinamycin 100%, gentamycin 100%, ampicillin 92.5% and cefalotin 85.2%) and to a glycopeptide antibiotic (vancomycin 100%). The Staphylococcus aureus strain had good sensitivity to antibiotics except gentamycin (100%) and kanamycin (100%). The enterobacteria tested were all sensitive to phenicols, carbapenems, cephalosporins and aminoglycosides. However, E. coli showed high resistance to tetracycline. The different methods showed low prevalences of C. trachomatis and N. gonorrhoeae, so comparisons Test RapidChlamydia/qPCR for C. trachomatis and culture/qPCR for N. gonorrhoeae were not possible. For S. agalactiae, on the other hand, qPCR was more advantageous than culture. The χ2 test showed a significant difference (Yates χ2 = 33.77 and p = 1-7) for the diagnosis of S. agalactiae. S. agalactiae qPCR had a sensitivity of 40.7%, a specificity of 94%, and positive and negative predictive values of 36.7% and 94.9% respectively, as well as a kappa = 0.33. The methods applied enabled us to identify the pathogens that cause cervicovaginal infections. The results suggest that qPCR may be an alternative, at least for the diagnosis of S. agalactiae. However, culture remains indispensable for studying antibiotic sensitivity. In order to improve patient care, molecular techniques need to be integrated into the HPD testing toolbox. To broaden the repertoire of pathogens to be diagnosed by qPCR, targeted comparison studies will be needed to increase the probability of encountering infected individuals.

Structural and biochemical analysis of penicillin-binding protein 2 from Campylobacter jejuni

Penicillin-binding protein 2 (PBP2) plays a key role in the formation of peptidoglycans in bacterial cell walls by crosslinking glycan chains through transpeptidase activity. PBP2 is also found in Campylobacter jejuni, a pathogenic bacterium that causes food-borne enteritis in humans. To elucidate the essential structural features of C. jejuni PBP2 (cjPBP2) that mediate its biological function, we determined the crystal structure of cjPBP2 and assessed its protein stability under various conditions. cjPBP2 adopts an elongated two-domain structure, consisting of a transpeptidase domain and a pedestal domain, and contains typical active site residues necessary for transpeptidase activity, as observed in other PBP2 proteins. Moreover, cjPBP2 responds to β-lactam antibiotics, including ampicillin, cefaclor, and cefmetazole, suggesting that β-lactam antibiotics inactivate cjPBP2. In contrast to typical PBP2 proteins, cjPBP2 is a rare example of a Zn2+-binding PBP2 protein, as the terminal structure of its transpeptidase domain accommodates a Zn2+ ion via three cysteine residues and one histidine residue. Zn2+ binding helps improve the protein stability of cjPBP2, providing opportunities to develop new C. jejuni-specific antibacterial drugs that counteract the Zn2+-binding ability of cjPBP2.

Rise in the Pathogenic Status of Coagulase-Negative Staphylococci Causing Bloodstream Infection.

Coagulase-negative staphylococci (CoNS) are one of the frequently isolated bacteria from blood cultures. Since they are part of the normal skin flora, they were previously considered contaminants. But now, they can be considered as established pathogens causing bloodstream infection (BSI). This study aims to estimate the prevalence of CoNS in BSI cases. This study was conducted at the Microbiology Department, All India Institute of Medical Sciences (AIIMS), Raipur, India, for eight months (January 2022 to August 2022). Data were collected retrospectively from medical and laboratory records. Paired blood cultures from 5085 clinically suspected sepsis cases were subjected to aerobic culture for five days in the BacT ALERT 3D system. Pathogenicity was established after recovery of CoNS from paired blood cultures of symptomatic patients. CoNS were isolated from 2.35% of patients, the most common species being Staphylococcus haemolyticus (51.67%). About 90% of isolates were methicillin-resistant. All the isolates were susceptible to linezolid, teicoplanin, and vancomycin, except one isolate of S. haemolyticus which was intermediate to vancomycin. Minimum inhibitory concentration (MIC) 50 and MIC 90 for vancomycin were 1 ug/ml and 2 ug/ml, respectively. Conclusion: Paired blood cultures are necessary to determine the pathogenicity of CoNS in BSI cases. A high prevalence of methicillin resistance, accompanied by high resistance rates to other non-beta lactam antibiotics, warrants the strict implementation of antimicrobial stewardship practices.

Synergism with Shikimic Acid Restores β-Lactam Antibiotic Activity against Methicillin-Resistant Staphylococcus aureus.

Methicillin-resistant Staphylococcus aureus (MRSA) has evolved into a dangerous pathogen resistant to beta-lactam antibiotics (BLAs) and has become a worrisome superbug. In this study, a strategy in which shikimic acid (SA), which has anti-inflammatory and antibacterial activity, is combined with BLAs to restart BLA activity was proposed for MRSA treatment. The synergistic effects of oxacillin combined with SA against oxacillin resistance in vitro and in vivo were investigated. The excellent synergistic effect of the oxacillin and SA combination was confirmed by performing the checkerboard assay, time-killing assay, live/dead bacterial cell viability assay, and assessing protein leakage. SEM showed that the cells in the control group had a regular, smooth, and intact surface. In contrast, oxacillin and SA or the combination treatment group exhibited different degrees of surface collapse. q-PCR indicated that the combination treatment group significantly inhibited the expression of the mecA gene. In vivo, we showed that the combination treatment increased the survival rate and decreased the bacterial load in mice. These results suggest that the combination of oxacillin with SA is considered an effective treatment option for MRSA, and the combination of SA with oxacillin in the treatment of MRSA is a novel strategy.

Tetracycline susceptibility in multidrug resistant Staphylococcus aureus isolated from clinical samples in tertiary care hospitals of Kathmandu, Nepal.

Staphylococcus aureus, being an opportunistic pathogen, is frequently associated with various infections in human. The clinical sector feels challenge regarding the management of multidrug resistant S. aureus. Besides the B-lactam antibiotics, Glycopeptides, Tetracycline, Quinolones, Macrolide-lincosamide streptogramin B family of antibiotics are also commonly prescribed to deal with such infections as an alternative drug. Therefore, this study was conducted to assess tetracycline susceptibility in multidrug resistant S. aureus in clinical samples. This study was conducted over the period of 15 months from March 2020–May 2021. The clinical samples were collected from the three different tertiary care hospitals of Kathmandu, Dirghayu Guru Hospital, Chabahil, Bhaktapur Cancer Hospital, Bhaktapur, and Shankarapur Hospital, Jorpati. The sample processing and laboratory work was carried out in the Microbiology Laboratory of Dirghyau Guru Hospital to determine the proportion of different phenotypes of MDR among S. aureus and their association with methicillin resistance. Two hundred sixty six isolates of S. aureus were included in the study. Methicillin resistance was detected by cefoxitin disc diffusion method. Among 2004 clinical samples processed, 266 (13.27%) isolates were S. aureus. The multidrug resistant strain isolated from OPD was 56.84% (54/95) and from inpatient was 43.15% (41/95). Among the 266 S. aureus, 28.94% MRSA and 71.05% MSSA isolates were detected. Tetracycline resistance was found in 9.8% of total S. aureus isolates whereas 27.36% (26/95) among MDR S. aureus isolates. Resistance to Tetracycline was higher in MRSA as compared to MSSA. Detection rate of MRSA in this study shows the necessity to improve health care practices to control MRSA infections. Tetracycline can be used as an alternative antibiotic against the S. aureus infection.

QUALITY CHARACTERISTICS OF RAW MILK SAMPLES PURCHASED FROM AUTOMATIC MİLK VENDING MACHINES IN NİĞDE PROVINCE

In the regulation published by the Ministry of Agriculture and Forestry in Turkey, it is stated that only automatic milk machines can be used to put raw milk on sale. This study aims to examine some physical, chemical and microbiological properties of a total of 40 raw milk samples sold in automatic vending machines and unpackaged, by egulations in Niğde province, and to compare the milk samples according to the retailer from which they were purchased. The study also aimed to quantitatively determine carbonate and hydrogen peroxide residues in milk samples and the presence of beta-lactam and tetracycline group antibiotics with a commercial kit. At the end of the study, the average value of lactic acid (%) density, fat, protein, non-fat dry matter, lactose, freezing point and pH values of the samples were determined as 0.153% ± (0.022); 1.028± (0.03) g/ml; 3.525%± (0.656); 3.5%± (0.107); 9% 4± (0.277); 5.134± (0.152); -0.549°C ± (0.018) and 6.55 ± (0.102) respectively. As a result of microbiological analysis of milk, the average number of total aerobic mesophilic organisms, coliforms, fecal coliforms, yeast-molds and micrococci-staphylococci was determined as 5.38 ± (0.47); 3.73 ± (1.11); 2.76 ± (1.66); 2.33 ± (1.86) and 4.29± (1.20) log cfu/ml respectively. Carbonate, hydrogen peroxide, beta-lactam and tetracycline antibiotic residues could not be detected in the milk samples. The fact that some of the results obtained in the study were outside the limits specified in the codexes and different from the study average reveals that the milk sold in businesses selling street milk should be analyzed regularly.

Effect of Saponin on Methylene Blue (MB) Photo-Antimicrobial Activity Against Planktonic and Biofilm Form of Bacteria.

Bacterial resistance has led to the spread of bacterial infections such as chronic wound infections. Finding solutions for combating resistant bacteria in chronic wounds such as Staphylococcus aureus and Pseudomonas aeruginosa became an attractive theme among researchers. P. aeruginosa is a gram negative opportunistic human pathogenic bacterium that is difficult to treat due to its high resistance to antibiotics. S. aureus (gram negative bacterium) also has a high antibiotic resistance, so that it is resistant to vancomycin (VRSA), tetracycline, fluoroquinolones and beta-lactam antibiotics including penicillin and methicillin (MRSA). In particular, S. aureus and P. aeruginosa have intrinsic and acquired antibiotic resistance, making the clinical management of infection a real challenge, especially in patients with comorbidities. aPDT can be proposed as a new method in the treatment of multi-drug resistant bacteria in chronic wound infection conditions. In this study, the effect of saponin (100 μg/mL) on photodynamic inactivation on planktonic and biofilm forms of P. aeruginosa (ATCC 27853) and S. aureus (ATCC 25923) strains and on Human Dermal Fibroblast (HDF) cells was investigated. Methylene blue (MB) was used as photosensitizer (0, 10, 50, 100 μg/mL). The light source was a red LED source (660 nm; power density: 20 mW/cm2) which is related to the maximum absorption of MB. The results showed that the use of saponin in combination with MB-aPDT (Methylene Blue-antibacterial photodynamic therapy) reduces the phototoxic activity of MB due to decreasing the monomer form of MB. This result was obtained by spectrophotometric study. Also, the result of MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide) assay showed that 8 min of irradiation (660 nm)at 10 μg/mL concentration of alone MB had the lowest phototoxic effect on HDF cells. Due to reduced phototoxic properties of MB in this method, detergents containing saponins not recommended to applied at the same time with MB-aPDT in wound infection area.

Structural and kinetic analysis of the monofunctional Staphylococcus aureus PBP1

Staphylococcus aureus, an ESKAPE pathogen, is a major clinical concern due to its pathogenicity and manifold antimicrobial resistance mechanisms. The commonly used β-lactam antibiotics target bacterial penicillin-binding proteins (PBPs) and inhibit crosslinking of peptidoglycan strands that comprise the bacterial cell wall mesh, initiating a cascade of effects leading to bacterial cell death. S. aureus PBP1 is involved in synthesis of the bacterial cell wall during division and its presence is essential for survival of both antibiotic susceptible and resistant S. aureus strains. Here, we present X-ray crystallographic data for S. aureus PBP1 in its apo form as well as acyl-enzyme structures with distinct classes of β-lactam antibiotics representing the penicillins, carbapenems, and cephalosporins, respectively: oxacillin, ertapenem and cephalexin. Our structural data suggest that the PBP1 active site is readily accessible for substrate, with little conformational change in key structural elements required for its covalent acylation of β-lactam inhibitors. Stopped-flow kinetic analysis and gel-based competition assays support the structural observations, with even the weakest performing β-lactams still having comparatively high acylation rates and affinities for PBP1. Our structural and kinetic analysis sheds insight into the ligand–PBP interactions that drive antibiotic efficacy against these historically useful antimicrobial targets and expands on current knowledge for future drug design and treatment of S. aureus infections.

Molecular characteristics and biofilm formation capacity of nontypeable Haemophilus influenza strains isolated from lower respiratory tract in children

With the widespread introduction of the Hib conjugate vaccine, Nontypeable Haemophilus influenzae (NTHi) has emerged as the predominant strain globally. NTHi presents a significant challenge as a causative agent of chronic clinical infections due to its high rates of drug resistance and biofilm formation. While current research on NTHi biofilms in children has primarily focused on upper respiratory diseases, investigations into lower respiratory sources remain limited. In this study, we collected 54 clinical strains of lower respiratory tract origin from children. Molecular information and drug resistance features were obtained through whole gene sequencing and the disk diffusion method, respectively. Additionally, an in vitro biofilm model was established. All clinical strains were identified as NTHi and demonstrated the ability to form biofilms in vitro. Based on scanning electron microscopy and crystal violet staining, the strains were categorized into weak and strong biofilm-forming groups. We explored the correlation between biofilm formation ability and drug resistance patterns, as well as clinical characteristics. Stronger biofilm formation was associated with a longer cough duration and a higher proportion of abnormal lung imaging findings. Frequent intake of β-lactam antibiotics might be associated with strong biofilm formation. While a complementary relationship between biofilm-forming capacity and drug resistance may exist, further comprehensive studies are warranted. This study confirms the in vitro biofilm formation of clinical NTHi strains and establishes correlations with clinical characteristics, offering valuable insights for combating NTHi infections.

β-lactam-induced OMV release promotes polymyxin tolerance in Salmonella enterica sv. Typhi.

The rise of multidrug-resistant bacteria is a global concern, leading to a renewed reliance on older antibiotics like polymyxins as a last resort. Polymyxins, cationic cyclic peptides synthesized nonribosomally, feature a hydrophobic acyl tail and positively charged residues. Their antimicrobial mechanism involves initial interaction with Gram-negative bacterial outer-membrane components through polar and hydrophobic interactions. Outer membrane vesicles (OMVs), nano-sized proteoliposomes secreted from the outer membrane of Gram-negative bacteria, play a crucial role in tolerating harmful molecules, including cationic peptides such as polymyxins. Existing literature has documented environmental changes' impact on modulating OMV properties in Salmonella Typhimurium. However, less information exists regarding OMV production and characteristics in Salmonella Typhi. A previous study in our laboratory showed that S. Typhi ΔmrcB, a mutant associated with penicillin-binding protein (PBP, a β-lactam antibiotic target), exhibited hypervesiculation. Consequently, this study investigated the potential impact of β-lactam antibiotics on promoting polymyxin tolerance via OMVs in S. Typhi. Our results demonstrated that sub-lethal doses of β-lactams increased bacterial survival against polymyxin B in S. Typhi. This phenomenon stems from β-lactam antibiotics inducing hypervesiculation of OMVs with higher affinity for polymyxin B, capturing and diminishing its biologically effective concentration. These findings suggest that β-lactam antibiotic use may inadvertently contribute to decreased polymyxin effectivity against S. Typhi or other Gram-negative bacteria, complicating the effective treatment of infections caused by these pathogens. This study emphasizes the importance of evaluating the influence of β-lactam antibiotics on the interaction between OMVs and other antimicrobial agents.

Metallo-β-lactamase inhibitors: A continuing challenge for combating antibiotic resistance

β-lactam antibiotics are the most successful and commonly used antibacterial agents, but the emergence of resistance to these drugs has become a global health threat. The expression of β-lactamase enzymes produced by pathogens, which hydrolyze the amide bond of the β-lactam ring, is the major mechanism for bacterial resistance to β-lactams. In particular, among class A, B, C and D β-lactamases, metallo-β-lactamases (MBLs, class B β-lactamases) are considered crucial contributors to resistance in gram-negative bacteria. To combat β-lactamase-mediated resistance, great efforts have been made to develop β-lactamase inhibitors that restore the activity of β-lactams. Some β-lactamase inhibitors, such as diazabicyclooctanes (DBOs) and boronic acid derivatives, have also been approved by the FDA. Inhibitors used in the clinic can inactivate mostly serine-β-lactamases (SBLs, class A, C, and D β-lactamases) but have not been effective against MBLs until now. In order to develop new inhibitors particularly for MBLs, various attempts have been suggested. Based on structural and mechanical studies of MBL enzymes, several MBL inhibitor candidates, including taniborbactam in phase 3 and xeruborbactam in phase 1, have been introduced in recent years. However, designing potent inhibitors that are effective against all subclasses of MBLs is still extremely challenging. This review summarizes not only the types of β-lactamase and mechanisms by which β-lactam antibiotics are inactivated, but also the research finding on β-lactamase inhibitors targeting these enzymes. These detailed information on β-lactamases and their inhibitors could give valuable information for novel β-lactamase inhibitors design.

Allergia agli antibiotici beta-lattamici: i miti da sfatare e le azioni pratiche da adottare

The penicillins (amoxicillin) are among the most effective and safe antibiotics for many common bacterial respiratory infections in paediatric age and should be avoided only when a true allergy is highly suspected. Severe true antibiotic allergies are rare and allergies are often overestimated. In high-income countries, 5-15% of patients report a penicillin allergy. However, in most cases (> 95%), these patients do not have a true immunologically mediated allergy and they may very likely tolerate the antibiotic upon a new exposure. All patients defined as allergic should be carefully evaluated and their levels of antibiotic allergy risk determined. In cases of suspected penicillin allergy (e.g. those with cutaneous manifestations), skin tests are not necessary before prescribing a beta-lactam antibiotic (amoxicillin) and direct oral administration can be performed in low-risk phenotypes carefully selected according to simple protocols that do not always require a specialist evaluation. The article, based on the recommendations of the WHO Aware Manual, provides current evidence and practical guidance, and fosters a correct interpretation and management of an overestimated problem that does not promote an optimal and judicious antibiotic use.

Full-dose challenge of moderate, severe, and unknown beta-lactam allergies in the emergency department.

This study aims to assess the outcome of challenging documented moderate, severe, or unknown beta-lactam allergies with full dose administration of a beta-lactam antibiotic in emergency department (ED) patients admitted for acute bacterial infection. A single-center, retrospective, descriptive study of adult patients challenged with a full dose of beta-lactam in the ED from January 2021 to December 2022 was conducted. Included patients had at least one documented moderate, severe, or unknown beta-lactam allergy in the electronic medical record (EMR) without documentation of prior tolerance. Patient demographics, prior beta-lactam antibiotic reaction, beta-lactam administered in the ED, inpatient beta-lactam continuation, adverse drug reactions, and updates to allergy profiles were collected. Descriptive statistics for data analysis were performed using SPSS Version 22. Of the 184 ED encounters with full-dose beta-lactam challenges, five (2.7%) patients with documented moderate, severe, or unknown beta-lactam allergies experienced an allergic reaction after the challenge; one (0.5%) patient had an allergic reaction in the ED, and the remaining four (2.2%) occurred after admission. No anaphylactic reactions occurred. All allergic reactions were limited to mild rash or itching. Most patients (98.9%) were challenged with a cephalosporin. A beta-lactam was continued in 86.4% of cases, and the allergy profile was updated for future utilization in 73.4% of patients. This study suggests that full-dose challenge of moderate, severe, or unknown beta-lactam allergies can be safely accomplished in the ED. This approach avoids unnecessary penicillin allergy skin testing and reduces utilization of suboptimal alternative antibiotic regimens.