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 ß-lactam antibiotics, including ampicillin, cefaclor, and cefmetazole, suggesting that ß-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.

Crucifer Lesion-Associated Xanthomonas Strains Show Multi-Resistance to Heavy Metals and Antibiotics.

Copper resistance in phytopathogens is a major challenge to crop production globally and is known to be driven by excessive use of copper-based pesticides. However, recent studies have shown co-selection of multiple heavy metal and antibiotic resistance genes in bacteria exposed to heavy metal and xenobiotics, which may impact the epidemiology of plant, animal, and human diseases. In this study, multi-resistance to heavy metals and antibiotics were evaluated in local Xanthomonas campestris pv. campestris (Xcc) and co-isolated Xanthomonas melonis (Xmel) strains from infected crucifer plants in Trinidad. Resistance to cobalt, cadmium, zinc, copper, and arsenic (V) was observed in both Xanthomonas species up to 25 mM. Heavy metal resistance (HMR) genes were found on a small plasmid-derived locus with ~ 90% similarity to a Stenotrophomonas spp. chromosomal locus and a X. perforans pLH3.1 plasmid. The co-occurrence of mobile elements in these regions implies their organization on a composite transposon-like structure. HMR genes in Xcc strains showed the lowest similarity to references, and the cus and ars operons appear to be unique among Xanthomonads. Overall, the similarity of HMR genes to Stenotrophomonas sp. chromosomal genomes suggest their origin in this genus or a related organism and subsequent spread through lateral gene transfer events. Further resistome characterization revealed the presence of small multidrug resistance (SMR), multidrug resistance (MDR) efflux pumps, and bla (Xcc) genes for broad biocide resistance in both species. Concurrently, resistance to antibiotics (streptomycin, kanamycin, tetracycline, chloramphenicol, and ampicillin) up to 1000 µg/mL was confirmed.

Distribution of bacteria and antimicrobial resistance in retail Nile tilapia (Oreochromis spp.) as potential sources of foodborne illness.

This study aimed to investigate AMR profiles of Aeromonas hydrophila, Salmonella spp., and Vibrio cholerae isolated from Nile tilapia (Oreochromis spp.) (n = 276) purchased from fresh markets and supermarkets in Bangkok, Thailand. A sample of tilapia was divided into three parts: fish intestine (n = 276), fish meat (n = 276), and liver and kidney (n = 276). The occurrence of A. hydrophila, Salmonella, and V. cholerae was 3.1%, 7.4%, and 8.5%, respectively. A high prevalence of these pathogenic bacteria was observed in fresh market tilapia compared to those from supermarkets (p < 0.05). The predominant Salmonella serovars were Paratyphi B (6.4%), followed by Escanaba (5.7%), and Saintpaul (5.7%). All isolates tested positive for the virulence genes of A. hydrophila (aero and hly), Salmonella (invA), and V. cholerae (hlyA). A. hydrophila (65.4%), Salmonella (31.2%), and V. cholerae (2.9%) showed multidrug resistant isolates. All A. hydrophila isolates (n = 26) exhibited resistant to ampicillin (100.0%) and florfenicol (100.0%), and often carried sul1 (53.8%) and tetA (50.0%). Salmonella isolates were primarily resistant to ampicillin (36.9%), with a high incidence of blaTEM (26.2%) and qnrS (25.5%). For V. cholerae isolates, resistance was observed against ampicillin (48.6%), and they commonly carried qnrS (24.3%) and tetA (22.9%). To identify mutations in the quinolone resistance determining regions (QRDRs), a single C248A point mutation of C248A (Ser-83-Tyr) in the gyrA region was identified in six out of seven isolates of Salmonella isolates. This study highlighted the presence of antimicrobial-resistant pathogenic bacteria in Nile tilapia at a selling point. It is important to rigorously implement strategies for AMR control and prevention.

Exploring ex vivo biofilm dynamics: consequences of low ampicillin concentrations on the human oral microbiome.

Prolonged exposure to antibiotics at low concentration can promote processes associated with bacterial biofilm formation, virulence and antibiotic resistance. This can be of high relevance in microbial communities like the oral microbiome, where commensals and pathogens share a common habitat and where the total abundance of antibiotic resistance genes surpasses the abundance in the gut. Here, we used an ex vivo model of human oral biofilms to investigate the impact of ampicillin on biofilm viability. The ecological impact on the microbiome and resistome was investigated using shotgun metagenomics. The results showed that low concentrations promoted significant shifts in microbial taxonomic profile and could enhance biofilm viability by up to 1 to 2-log. For the resistome, low concentrations had no significant impact on antibiotic resistance gene (ARG) diversity, while ARG abundance decreased by up to 84%. A positive correlation was observed between reduced microbial diversity and reduced ARG abundance. The WHO priority pathogens Streptococcus pneumoniae and Staphylococcus aureus were identified in some of the samples, but their abundance was not significantly altered by ampicillin. Most of the antibiotic resistance genes that increased in abundance in the ampicillin group were associated with streptococci, including Streptococcus mitis, a well-known potential donor of ARGs to S. pneumoniae. Overall, the results highlight the potential of using the model to further our understanding of ecological and evolutionary forces driving antimicrobial resistance in oral microbiomes.

Antimicrobial Resistance Patterns in Patients with Vaginal Discharge: A 2019-2022 Analysis at the National Health Laboratory in Eritrea.

Background: Antimicrobial resistance poses a significant global threat to the treatment of bacterial infections, particularly in low- and middle-income regions such as Africa. This study is aimed at analyzing antimicrobial resistance patterns in vaginal swab samples from patients at the National Health Laboratory from 2019 to 2022. Methods: This retrospective study examined patient records from vaginal swab analyses performed at the National Health Laboratory between January 1, 2019, and December 31, 2022. Ethical approval was obtained from the Ministry of Health Research Ethical Approval and Clearance Committee on 15/02/2023. Results: Of the 622 samples, 83% underwent microbial isolation and identification. Citrobacter spp. exhibited high resistance (>43%) to antibiotics such as cephalexin, ceftazidime, nalidixic acid, ampicillin, gentamicin, and tetracycline. E. coli showed resistance rates of more than 50% to ampicillin, trimethoprim-sulfamethoxazole, and tetracycline. Klebsiella spp. and Proteus spp. exhibited resistance rates that exceeded 47% to specific antibiotics. Gram-positive bacteria have resistance rates of more than 49% with ampicillin, trimethoprim-sulfamethoxazole, tetracycline, oxacillin, vancomycin, and penicillin G. In particular, S. aureus demonstrated no resistance to rifampicin or clindamycin, while Streptococcus spp. showed 100% resistance to rifampicin and vancomycin. Several species, including Proteus species, Streptococcus spp., S. aureus, and Klebsiella spp. exhibited multidrug resistance. Conclusion: Most gram-negative bacteria displayed higher resistance of >45% to ampicillin, trimethoprim-sulfamethoxazole, and tetracycline. Among gram-positive bacteria, a higher resistance rate with ampicillin, trimethoprim-sulfamethoxazole, tetracycline, oxacillin, vancomycin, and penicillin G was recorded. S. aureus showed no resistance to rifampicin and clindamycin, and Strep. spp. indicated 100% resistance to rifampicin and vancomycin. This study highlights critical gaps and areas for further exploration. Expanding the spectrum of antibiotics tested and investigating underlying multidrug resistance mechanisms would provide a more comprehensive understanding of resistance patterns.

Occurrence, antimicrobial susceptibility, and resistance genes of Staphylococcus aureus in milk and milk products in the Arsi highlands of Ethiopia.

BACKGROUND: In Ethiopia, milk production and handling practices often lack proper hygiene measures, leading to the potential contamination of milk and milk products with Staphylococcus aureus (S. aureus), including methicillin-resistant strains, posing significant public health concerns. This study aimed to investigate the occurrence, antimicrobial susceptibility profiles and presence of resistance genes in S. aureus strains isolated from milk and milk products. METHODS: A cross-sectional study was conducted in the Arsi highlands, Oromia, Ethiopia from March 2022 to February 2023. A total of 503 milk and milk product samples were collected, comprising 259 raw milk, 219 cottage cheese, and 25 traditional yogurt samples. S. aureus isolation and coagulase-positive staphylococci enumeration were performed using Baird-Parker agar supplemented with tellurite and egg yolk. S. aureus was further characterized based on colony morphology, Gram stain, mannitol fermentation, catalase test, and coagulase test. Phenotypic antimicrobial resistance was assessed using the Kirby-Bauer disc diffusion method, while the polymerase chain reaction (PCR) was employed for confirming the presence of S. aureus and detecting antimicrobial resistance genes. RESULTS: S. aureus was detected in 24.9% of the milk and milk products, with the highest occurrence in raw milk (40.9%), followed by yogurt (20%), and cottage cheese (6.4%). The geometric mean for coagulase-positive staphylococci counts in raw milk, yogurt, and cottage cheese was 4.6, 3.8, and 3.2 log10 CFU/mL, respectively. Antimicrobial resistance analysis revealed high levels of resistance to ampicillin (89.7%) and penicillin G (87.2%), with 71.8% of the isolates demonstrating multidrug resistance. Of the 16 S. aureus isolates analyzed using PCR, all were found to carry the nuc gene, with the mecA and blaZ genes detected in 50% of these isolates each. CONCLUSION: This study revealed the widespread distribution of S. aureus in milk and milk products in the Arsi highlands of Ethiopia. The isolates displayed high resistance to ampicillin and penicillin, with a concerning level of multidrug resistance. The detection of the mecA and blaZ genes in selected isolates is of particular concern, highlighting a potential public health hazard and posing a challenge to effective antimicrobial treatment. These findings highlight the urgent need to enhance hygiene standards in milk and milk product handling and promote the rational use of antimicrobial drugs. Provision of adequate training for all individuals involved in the dairy sector can help minimize contamination. These measures are crucial in addressing the threats posed by S. aureus, including methicillin-resistant strains, and ensuring the safety of milk and its products for consumers.

Rapid evolutionary change in trait correlations of single proteins.

Many organismal traits are genetically determined and covary in evolving populations. The resulting trait correlations can either help or hinder evolvability - the ability to bring forth new and adaptive phenotypes. The evolution of evolvability requires that trait correlations themselves must be able to evolve, but we know little about this ability. To learn more about it, we here study two evolvable systems, a yellow fluorescent protein and the antibiotic resistance protein VIM-2 metallo beta-lactamase. We consider two traits in the fluorescent protein, namely the ability to emit yellow and green light, and three traits in our enzyme, namely the resistance against ampicillin, cefotaxime, and meropenem. We show that correlations between these traits can evolve rapidly through both mutation and selection on short evolutionary time scales. In addition, we show that these correlations are driven by a protein's ability to fold, because single mutations that alter foldability can dramatically change trait correlations. Since foldability is important for most proteins and their traits, mutations affecting protein folding may alter trait correlations mediated by many other proteins. Thus, mutations that affect protein foldability may also help shape the correlations of complex traits that are affected by hundreds of proteins.

Epidemiología molecular de la colonización nasofaríngea neumocócica en niños de Sevilla, tras la implementación del programa de vacunación con VNC13 en Andalucía (España) / Molecular epidemiology of pneumococcal carriage in children from Seville, following implementation of the PCV13 immunization program in Andalusia, Spain

Introducción: El programa de vacunación universal con la vacuna antineumocócica conjugada 13-valente (VNC13) se implantó en Andalucía en diciembre de 2016. Métodos: Estudio transversal de colonización nasofaríngea por Streptococcus pneumoniae. Se seleccionó a 397 niños sanos en centros de atención primaria de Sevilla durante los periodos 1/4/2018-28/2/2020 y 1/11/2021-28/2/2022 (periodo VNC13). Se utilizó una colección histórica de un estudio de colonización desarrollado en niños sanos y con infección respiratoria superior entre el 1/01/2006 y el 30/06/2008 (periodo VNC7) para comparar las distribuciones de serotipos/genotipos y las tasas de resistencias antibióticas. Resultados: Un total de 76 (19%) niños estaban colonizados con S. pneumoniae en el periodo VNC13 y se dispuso de 154 aislamientos del periodo VNC7. La colonización por serotipos incluidos en VNC13 disminuyó significativamente entre los periodos VNC13 y VNC7 (11 vs. 38%; p=0,0001); los serotipos 19F (8%), 3 (1%) y 6B (1%) fueron los únicos serotipos vacunales circulantes. Los serotipos 15B/C y 11A fueron los serotipos no VNC13 más prevalentes durante el periodo VNC13 (14% y 11%, respectivamente); este último se incrementó de forma significativa entre periodos de tiempo (p=0,04). El serotipo 11A solo se asoció en el periodo VNC13 con variantes resistentes a la ampicilina del clon Spain9V-ST156 (ST6521 y genéticamente relacionado ST14698), no detectados en el periodo anterior. Conclusiones: Hubo una circulación muy residual de los serotipos vacunales durante el periodo VNC13, con excepción del serotipo19F. El serotipo 11A se incrementó de forma significativa entre los periodos VNC13 y VNC7 por expansión clonal del genotipo resistente a la ampicilina ST6521.(AU)

Background: The 13-valent pneumococcal conjugate vaccine (PCV13) universal vaccination program was introduced in December 2016 in Andalusia. Methods: A cross-sectional study was conducted on the molecular epidemiology of pneumococcal nasopharyngeal colonization. A total of 397 healthy children were recruited from primary healthcare centres in Seville for the periods 1/4/2018 to 28/2/2020 and 1/11/2021 to 28/2/2022 (PCV13 period). Data from a previous carriage study conducted among healthy and sick children from 1/01/2006 to 30/06/2008 (PCV7 period) were used for comparison of serotype/genotype distributions and antibiotic resistance rates. Results: Overall, 76 (19%) children were colonized with S. pneumoniae during the PCV13 period and there were information available from 154 isolates collected during the PCV7 period. Colonization with PCV13 serotypes declined significantly in the PCV13 period compared with historical controls (11 vs. 38%, P=0.0001), being serotypes 19F (8%), 3 (1%) and 6B (1%) the only circulating vaccine types. Serotypes 15B/C and 11A were the most frequently identified non-PCV13 serotypes during the PCV13 period (14% and 11%, respectively); the later one increased significantly between time periods (P=0.04). Serotype 11A was exclusively associated in the PCV13 period with ampicillin-resistant variants of the Spain9V-ST156 clone (ST6521 and genetically related ST14698), not detected in the preceding period. Conclusions: There was a residual circulation of vaccine types following PCV13 introduction, apart from serotype 19F. Serotype 11A increased between PCV13 and PCV7 periods due to emergence and clonal expansion of ampicillin-resistant genotype ST6521.(AU)

Listeria monocytogenes: the silent assassin.

Listeriosis is a foodborne infection in humans caused by Listeria monocytogenes. Consumption of contaminated food can lead to severe infection in vulnerable patients, that can be fatal. Clinical manifestations include sepsis and meningitis, and in pregnancy-associated infection, miscarriage and stillbirth. Diagnosis is confirmed by culture and identification of the pathogen from blood, cerebrospinal fluid, vaginal swab, placenta or amniotic fluid. Treatment regimens recommend amoxicillin, ampicillin or an aminoglycoside. Virulence factors mediate bacterial adhesion and invasion of gut epithelial cells. Other factors mediate biofilm formation and tolerance to low temperatures and high salt concentrations facilitating persistence and survival in the environment.

Prevalence and antimicrobial resistance of Enterobacteriaceae in the north of Kazakhstan.

Background: An increasing number of drugs are used each year in the treatment of small pets (cats and dogs), including medicines (cephalosporins and fluoroquinolones) used in human therapy. Aim: The purpose of this study was to isolate and explore the antibiotic resistance of opportunistic Enterobacteriaceae (Escherichia coli, Klebsiella, Proteus, Ci trobacter, Enterobacter) from cats and dogs, and to isolate resistance genes in the microorganisms. Methods: In 2021, 808 samples of biological material from small domestic animals were collected in veterinary clinics in Kostanay. From these, 210 microorganisms were isolated and identified. Results: A large majority of the strains sampled belonged to E. coli-149 (70.9%), Enterobacter-11 (5.2%), Klebsiella-28 (13.3%), Proteus-12 (5.7%) and 10 Citrobacter isolates (4.8%). In all isolates identified, antibiotic resistance/sensitivity was determined by disc-diffusion method to ampicillin, cefoxitin, gentamicin, levomycetin, tetracycline, ciprofloxacin, norfloxacin, ofloxacin, cefoperazone, cefpodoxime, streptomycin, kanamycin, doxycycline, gemifloxacin, nalidixic acid, furazolidone, furadonine, amoxicillin, and enrofloxacin. Conclusion: The study has demonstrated that the greatest number of Enterobacteriaceae were sensitive to the action of meropenem, which belongs to the group of beta-lactam antibiotics; resistance was demonstrated against tetracycline, doxycycline, ampicillin, amoxicillin, ofloxacin, and cefpodoxime. The most common genes encoding antimicrobial resistance were as follows: BlaTEM and OXA in 41 and 28 isolates, respectively, encoding resistance to beta-lactams; StrA and StrB in 45 and 48 isolates encoding aminoglycosides; and tetA and tetB in 43 and 28 isolates encoding tetracyclines. Obtained data demonstrate that uncontrolled and frequent use of beta-lactam and tetracycline antibacterials, in cats and dogs, results in the spread of genotypic resistance among micro-organisms of the family Enterobacteriaceae.

Label-free electrochemical sensing platform for sensitive detection of ampicillin by combining nucleic acid isothermal enzyme-free amplification circuits with CRISPR/Cas12a.

The residue of ampicillin (AMP) in food and ecological environment poses a potential harm to human health. Therefore, a reliable system for detecting AMP is in great demand. Herein, a label-free and sensitive electrochemical sensor utilizing NH2-Co-MOF as an electrocatalytic active material for methylene blue (MB) was developed for rapid and facile AMP detection by combining hybridization chain reaction (HCR), catalytic hairpin assembly (CHA) with CRISPR/Cas12a. The surface of glassy carbon electrode modified with NH2-Co-MOF was able to undergo HCR independent of the AMP, forming long dsDNA complexes to load MB, resulting in strong original electrochemical signal. The presence of AMP could trigger upstream CHA circuit to activate the CRISPR/Cas12a system, thereby achieving rapid non-specific cleavage of the trigger ssDNA of HCR on the electrode surface, hindering the occurrence of HCR and reducing the load of MB. Significant signal change triggered by the target was ultimately obtained, thus achieving sensitive detection of the AMP with a detection limit as low as 1.60 pM (S/N = 3). The proposed sensor exhibited good stability, selectivity, and stability, and achieved reliable detection of AMP in milk and livestock wastewater samples, demonstrating its promising application prospects in food safety and environmental monitoring.

Mesoporous silica nanospheres-mediated insecticide and antibiotics co-delivery system for synergizing insecticidal toxicity and reducing environmental risk of insecticide.

Mesoporous silica nanoparticles (MSNs) are efficient carriers of drugs, and are promising in developing novel pesticide formulations. The cotton aphids Aphis gossypii Glover is a world devastating insect pest. It has evolved high level resistance to various insecticides thus resulted in the application of higher doses of insecticides, which raised environmental risk. In this study, the MSNs based pesticide/antibiotic delivery system was constructed for co-delivery of ampicillin (Amp) and imidacloprid (IMI). The IMI@Amp@MSNs complexes have improved toxicity against cotton aphids, and reduced acute toxicity to zebrafish. From the 16S rDNA sequencing results, Amp@MSNs, prepared by loading ampicillin to the mesoporous of MSNs, greatly disturbed the gut community of cotton aphids. Then, the relative expression of at least 25 cytochrome P450 genes of A. gossypii was significantly suppressed, including CYP6CY19 and CYP6CY22, which were found to be associated with imidacloprid resistance by RNAi. The bioassay results indicated that the synergy ratio of ampicillin to imidacloprid was 1.6, while Amp@MSNs improved the toxicity of imidacloprid by 2.4-fold. In addition, IMI@Amp@MSNs significantly improved the penetration of imidacloprid, and contributed to the amount of imidacloprid delivered to A. gossypii increased 1.4-fold. Thus, through inhibiting the relative expression of cytochrome P450 genes and improving penetration of imidacloprid, the toxicity of IMI@Amp@MSNs was 6.0-fold higher than that of imidacloprid. The greenhouse experiments further demonstrated the enhanced insecticidal activity of IMI@Amp@MSNs to A. gossypii. Meanwhile, the LC50 of IMI@Amp@MSNs to zebrafish was 3.9-fold higher than that of IMI, and the EC50 for malformation was 2.8-fold higher than IMI, respectively, which indicated that the IMI@Amp@MSNs complexes significantly reduced the environmental risk of imidacloprid. These findings encouraged the development of pesticide/antibiotic co-delivery nanoparticles, which would benefit pesticide reduction and environmental safety.

Degradation of veterinary antibiotics by the ozonation process: Product identification and ecotoxicity assessment.

The purpose of the study was to evaluate the effects of using of ozonation to remove antibiotics used, among others, in veterinary medicine, from the aqueous environment. The effect of this process on the degradation, mineralisation and ecotoxicity of aqueous solutions of ampicillin, doxycycline, tylosin, and sulfathiazole was investigated. Microbiological MARA® bioassay and two in silico methods were used for the ecotoxicity assessment. Ozonation was an effective method for the degradation of the antibiotics studied and the reduction in ecotoxicity of the solutions. However, after ozonation, the solutions contained large amounts of organic products, including compounds much less susceptible to ozonation than the initial antibiotics. Structures of 14, 12, 40 and 10 degradation products for ampicillin, doxycycline, tylosin, and sulfathiazole, respectively, were proposed. It was confirmed that ozone plays a greater role than hydroxyl radicals in the degradation of these antibiotics, with the exception of TYL. The use of ozonation to obtain a high degree of mineralisation is unfavourable and it is suggested to combine ozonation with biodegradation. The pre-ozonation will cause decomposition of antibiotic pharmacophores, which significantly reduces the risk of spread of antimicrobial resistance in the active biocenosis of wastewater treatment plants.

Comparative analysis of characteristics of antibiotic resistomes between Arctic soils and representative contaminated samples using metagenomic approaches.

Antibiotic resistance is one of the most concerned global health issues. However, comprehensive profiles of antibiotic resistance genes (ARGs) in various environmental settings are still needed to address modern antibiotic resistome. Here, Arctic soils and representative contaminated samples from ARG pollution sources were analyzed using metagenomic approaches. The diversity and abundance of ARGs in Arctic soils were significantly lower than those in contaminated samples (p < 0.01). ARG profiles in Arctic soils were featured with the dominance of vanF, ceoB, and bacA related to multidrug and bacitracin, whereas those from ARG pollution sources were characterized by prevalent resistance to anthropogenic antibiotics such as sulfonamides, tetracyclines, and beta-lactams. Mobile genetic elements (MGEs) were found in all samples, and their abundance and relatedness to ARGs were both lower in Arctic soils than in polluted samples. Significant relationships between bacterial communities and ARGs were observed (p < 0.01). Cultural bacteria in Arctic soils had clinically-concerned resistance to erythromycin, vancomycin, ampicillin, etc., but ARGs relevant to those antibiotics were undetectable in their genomes. Our results suggested that Arctic environment could be an important reservoir of novel ARGs, and antibiotic stresses could cause ARG pollution via horizontal gene transfer and enrichment of resistant bacteria.

Cell surface ß-lactamase recruitment: A facile selection to identify protein-protein interactions.

Protein-protein interactions (PPIs) are central to many cellular processes, and the identification of novel PPIs is a critical step in the discovery of protein therapeutics. Simple methods to identify naturally existing or laboratory evolved PPIs are therefore valuable research tools. We have developed a facile selection that links PPI-dependent ß-lactamase recruitment on the surface of Escherichia coli with resistance to ampicillin. Bacteria displaying a protein that forms a complex with a specific protein-ß-lactamase fusion are protected from ampicillin-dependent cell death. In contrast, bacteria that do not recruit ß-lactamase to the cell surface are killed by ampicillin. Given its simplicity and tunability, we anticipate this selection will be a valuable addition to the palette of methods for illuminating and interrogating PPIs.

Antibacterial efficacy, mode of action, and safety of a novel nano-antibiotic against antibiotic-resistant Escherichia coli strains.

Globally rising antibiotic-resistant (AR) and multi-drug resistant (MDR) bacterial infections are of public health concern due to treatment failure with current antibiotics. Enterobacteria, particularly Escherichia coli, cause infections of surgical wound, bloodstream, and urinary tract, including pneumonia and sepsis. Herein, we tested in vitro antibacterial efficacy, mode of action (MoA), and safety of novel amino-functionalized silver nanoparticles (NH2-AgNP) against the AR bacteria. Two AR E. coli strains (i.e., ampicillin- and kanamycin-resistant E. coli), including a susceptible strain of E. coli DH5α, were tested for susceptibility to NH2-AgNP using Kirby-Bauer disk diffusion and standard growth assays. Dynamic light scattering (DLS) was used to determine cell debris and relative conductance was used as a measure of cell leakage, and results were confirmed with transmission electron microscopy (TEM). Multiple oxidative stress assays were used for in vitro safety evaluation of NH2-AgNP in human lung epithelial cells. Results showed that ampicillin and kanamycin did not inhibit growth in either AR bacterial strain with doses up to 160 µg/mL tested. NH2-AgNP exhibited broad-spectrum bactericidal activity, inhibiting the growth of all three bacterial strains at doses ≥1 µg/mL. DLS and TEM revealed cell debris formation and cell leakage upon NH2-AgNP treatment, suggesting two possible MoAs: electrostatic interactions followed by cell wall damage. Safety evaluation revealed NH2-AgNP as noncytotoxic and antioxidative to human lung epithelial cells. Taken together, these results suggest that NH2-AgNP may serve as an effective and safer bactericidal therapy against AR bacterial infections compared to common antibiotics.

Clinical characteristics and risk factors for Klebsiella pneumoniae bloodstream infection in 152 immunocompetent patients.

OBJECTIVE: To investigate the clinical characteristics and prognostic risk factors for Klebsiella pneumoniae bloodstream infections in immunocompetent patients. METHODS: The study included patients with K. pneumoniae bloodstream infection treated in Zhongda Hospital from June 2016 to June 2021. Clinical data and antibiotic susceptibility test results were retrospectively collected and analyzed. Independent risk factors for mortality were screened using the chi-square test and multivariate logistic regression. RESULTS: A total of 152 patients were included in the analysis. In our cohort, 77.6% of patients were older than 60 years, and 80.9% of them had community-acquired infections. The most common complications were type 2 diabetes, hypertension, and stroke sequelae. The proportion of patients with septic shock or abscesses was 34.9% and 25.7%, respectively. There were significant differences in the site of infection, septic shock, and serum levels of procalcitonin, hypersensitive C-reactive protein, D-dimer, creatinine, and lactic acid between survivors and non-survivors (p < 0.05). Multivariate regression analysis showed that hospital-acquired infections, septic shock, length of hospital stay, and creatinine levels were independent risk factors for mortality. Antibiotic susceptibility test results indicated that clinical outcomes varied depending on bacterial sensitivity to ampicillin/sulbactam. DISCUSSION: Klebsiella pneumoniae is a common community-acquired and hospital-acquired bacteria and usually infects older people with complications such as diabetes. Nosocomial infections, length of stay, septic shock, and renal insufficiency are potentially associated with poor prognosis. Bacterial susceptibility to ampicillin/sulbactam affects prognosis.

Source tracking of extensively drug resistant Salmonella Typhi in food and raw vegetables using molecular approaches.

INTRODUCTION: Extensively drug resistant (XDR) strains of the Salmonella lineages have been reported to spread from Africa to South Asia. XDR strains are resistant to fluoroquinolones, chloramphenicol, co-trimoxazole, and ampicillin, resulting in treatment failure. The objectives of this study included the investigation of transmission of S. Typhi lineages and the identification of the potentially contaminated sources of the XDR typhoid outbreak from different urban areas by using molecular techniques. METHODOLOGY: Environmental samples, including food samples, were collected from different towns and the susceptibility of each isolate to the antimicrobial agents was examined. Molecular identification of different Salmonella lineages including S. Typhi, S. Paratyphi A, H58, and XDR was carried out through multiplex PCR. RESULTS AND CONCLUSIONS: A total of 328 environmental samples including raw vegetables, water, and bakery items were collected. More than half of the tested samples (64%) found harboring Salmonella spp. The Salmonella was confirmed through PCR amplification of species-specific markers that showed the presence of S. Typhi (40%), S. Paratyphi A (8%), H58 (7%), and XDR S. Typhi (6%). Raw vegetables had the highest number of Salmonella spp., indicating consumption of raw vegetables as a possible source of salmonellosis. XDR status was also affirmed through phenotypic antimicrobial susceptibility testing.

Antimicrobial resistance profiles of Pseudomonas aeruginosa, Escherichia coli and Klebsiella pneumoniae strains isolated from broiler chickens.

Globally, the spread of multidrug-resistant Pseudomonas aeruginosa, Escherichia coli, and Klebsiella pneumoniae from food to humans poses a severe threat to public health. The aim of this study was to assess the co-occurrence of colistin and ß-lactamase resistance genes in E. coli, K. pneumoniae, and P. aeruginosa strains isolated from faeces of abattoir broiler chickens. The E. coli, P. aeruginosa and K. pneumoniae isolates were successfully detected from faecal samples by polymerase chain reaction (PCR) at infection rates of 60.7%, 22.5% and 16.7% respectively. The isolates displayed the highest levels of antibiotic resistance (AR) against ampicillin (82.3%) and amoxicillin-clavulanic acid (74.2%) for E. coli, followed by cefoxitin (70.6%) for K. pneumoniae, whilst P. aeruginosa displayed 26.1% antibiotic resistance (AR) against both ampicillin and colistin sulphate. The colistin mcr-1 gene was harboured by 46.8%, 47.1% and 21.7%, E. coli, K. pneumonia and P. aeruginosa isolates respectively. Ten out of 62 (16.1%), 6/17 (35.3%), 4/23 (17.4%) isolates were phenotypically classified as ESBL E. coli, K. pneumoniae, and P. aeruginosa respectively. The ESBL-E. coli isolates respectively possessed blaCTX-M (60%), blaTEM (20%) and blaCTX-M-9 (10%) genes. The ESBL-K. pneumoniae harboured, blaCTX-M (50%), blaOXA (33%), blaCARB (17%), and blaCTX-M-9 (17%) genes respectively, whilst, P. aeruginosa isolates respectively carried blaTEM (75%), blaCTX-M (50%), blaOXA (25%) and blaCARB (25%) genes. Molecular analysis identified the blaCTX-Mß-lactamase-encoding genes collectively from E. coli, P. aeruginosa, K. pneumoniae isolates. Colistin and ß-lactamase genes were present in only 16.7%, 6.9%, and 2.9% of E. coli, K. pneumoniae, and P. aeruginosa isolates, respectively. A total of 17, 7 and 3 isolates for E. coli, K. pneumoniae and P. aeruginosa respectively carried both colistin and ß-lactamase antibiotics resistant genes. This is a public health threat that points to a challenge in the treatment of infections caused by these zoonotic bacteria. Data generated from this study will contribute to formulation of new strategies for combating spread of E. coli, K. pneumoniae, and P. aeruginosa isolates as well as prevention of their AR development.

[Virulence genes and antimicrobial resistance of Bacillus cereus in prepackaged pastries sold in Taizhou from 2020 to 2022].

OBJECTIVE: To investigate the virulence genes and antimicrobial resistance of Bacillus cereus from the pre-packaged pastries in Taizhou city. METHODS: 500 pre-packaged patries were collected in taizhou city market. 97 Bacillus cereus strains were detected from them by GB 4789.14-2014 method and identified with 4 houseking genes, then 13 virulence genes were detected by polymerase chain reaction(PCR)method and the antimicrobial resistance of Bacillus cereus to 19 antibiotics was detected by paper diffusion method. RESULTS: The result showed that the contamination rate of Bacillus cereus was 19.4% in 500 pre-packaged pastries. The detection rate of four housekeeping genes groEL, gyr B, rpoB and Vrr were 100%, 94.8%, 97.9% and 96.9%, respectively, and 89.7% at the same time. The virulence gene test result showed that the detection rate of nheABC, entFM, bceT, cytK and hblABCD were 91.8%, 88.7%, 61.9%, 51.6% and 25.8%, emetic virulence genes had the lowest detection rate, ces and EMl were 4.1%, cer was 5.2%. 97 Bacillus cereus strains show different degrees of drug resistance to 14 antimicrobials, the resistance rates to penicillin, ampicillin, cefotaxime and cotrimoxazole were higher than 95%, but they were completely sensitive to streptomycin, vancomycin and chloramphenicol. CONCLUSION: There is a risk of contamination by diarrhea-type Bacillus cereus and vomiting-type Bacillus cereus in prepackaged pastries in Taizhou. The isolated and identified Bacillus cereus has multiple-drug resistance.