Methicillin-resistant Research Articles

Isolation and molecular characterization of multi-drug resistant uropathogenic Escherichia coli from urine samples: Insights into urinary tract infection management

Urinary tract infections (UTIs) caused by uropathogenic Escherichia coli (UPEC) present a significant global health burden. With UPEC responsible for approximately 90 % of UTIs, understanding its mechanisms of virulence and antibiotic resistance is paramount for effective treatment strategies. This study aimed to investigate the prevalence of multi-drug resistant (MDR) E. coli strains in urinary tract infections and to elucidate the molecular mechanisms underlying their resistance. A total of 150 E. coli isolates were obtained from urinary tract infection specimens collected patients at Doctor’s Diagnostic Centre in Kalyan Nagar, Bangalore over four months November 2022 to February 2023. Isolation and screening of pathogenic E. coli were conducted using Eosin Methylene Blue Agar and Nutrient Agar, followed by identification through gram staining and biochemical characterization tests. Antibiotic susceptibility testing was performed using Mueller-Hinton agar and various antibiotic discs. Phenotypic and molecular identification of extended-spectrum beta-lactamase (ESBL) producers were conducted, along with the screening for specific antibiotic resistance genes using polymerase chain reaction (PCR). Of the 150 E. coli isolates, 57.3 % were found to be multi-drug resistant, with all 35 isolates from hospitalized patients exhibiting MDR characteristics. Phenotypic and molecular analyses revealed the presence of ESBL-producing isolates, with the blaCTX-M, blaTEM, and blaSHV genes detected in PCR assays. Plasmid extraction and PCR assays further identified the presence of plasmid-encoded carbapenemase genes (blaOXA-48, blaNDM-1, blaKPC-2), as well as other antibiotic resistance genes including those conferring resistance to fluoroquinolones. The results underscore the alarming prevalence of MDR E. coli strains in urinary tract infections, particularly in hospitalized patients. Molecular characterization revealed the diverse genetic mechanisms contributing to antibiotic resistance, including the presence of ESBLs and plasmid-encoded resistance genes. These findings emphasize the urgent need for surveillance and infection control measures to mitigate the spread of MDR pathogens and the development of alternative therapeutic strategies to combat antibiotic resistance in UTIs.

Intensive care unit admission rates and factors associated following Autologous stem cell transplantation-real-world experience from a tertiary center in rural India.

Infectious and other complications can necessitate admission to the intensive care unit (ICU) in autologous stem cell transplantation (ASCT). Data on need for ICU care, impact of various pre- and peri-transplant characteristics on requirement of ICU care and outcomes are scarce from the developing world. A retrospective case record review of ASCT cases was conducted. Pre- and peri-transplant characteristics like infection within 4weeks of transplant, mucositis, surveillance culture positivity, peri-transplant infections, comorbidity, and time to neutrophil and platelet engraftment were noted. A total of 109 patients underwent 109 ASCTs. Most common diagnosis was the plasma cell disorder in 75 (69%) patients. Forty-eight (45%) patients had peri-transplant infections. Fifteen (14%) patients had infections with multi-drug resistant (MDR) organisms. Fifteen (14%) patients required ICU care, the most common reason being hypotension in nine patients (8.3%). Four patients (3.7%) required non-invasive ventilation, and one (0.9%) required invasive ventilation. Mortality rate was 1.8% (two patients). Factors associated with the need for ICU care were time to platelet engraftment (median 15days among those required ICU care versus 13days who did not, p = 0.04) and presence of peri-transplant infection showed a trend toward ICU care need (19% among those required ICU care versus 7% in those who did not, p = 0.05). Delayed platelet engraftment was associated with the need for ICU care and peri-transplant infections were associated with a trend toward need for ICU care.

Green synthesis of cerium oxide nanoparticles usingTribulus terrestris: characterization and evaluation of antioxidant, anti-inflammatory and antibacterial efficacy against wound isolates.

Multi-drug resistance (MDR) infections are a significant global challenge, necessitating innovative and eco-friendly approaches for developing effective antimicrobial agents. This study focuses on the synthesis, characterization, and evaluation of cerium oxide nanoparticles (CeO2NPs) for their antioxidant, anti-inflammatory, and antibacterial properties. The CeO2NPs were synthesized using aTribulus terrestrisaqueous extract through an environmentally friendly process. Characterization techniques included UV-visible spectroscopy, Fourier Transform Infrared Spectroscopy (FT-IR), x-ray Diffraction (XRD), Scanning Electron Microscopy (SEM), and Energy Dispersive x-ray (EDX) analysis. The UV-vis spectroscopy shows the presence of peak at 320 nm which confirms the formation of CeO2NPs. The FT-IR analysis of the CeO2NPs revealed several distinct functional groups, with peak values at 3287, 2920, 2340, 1640, 1538, 1066, 714, and 574 cm-1. These peaks correspond to specific functional groups, including C-H stretching in alkynes and alkanes, C=C=O, C=C, alkanes, C-O-C, C-Cl, and C-Br, indicating the presence of diverse chemical bonds within the CeO2NPs. XRD revealed that the nanoparticles were highly crystalline with a face-centered cubic structure, and SEM images showed irregularly shaped, agglomerated particles ranging from 100-150 nm. In terms of biological activity, the synthesized CeO2NPs demonstrated significant antioxidant and anti-inflammatory properties. The nanoparticles exhibited 82.54% antioxidant activity at 100 μg ml-1, closely matching the 83.1% activity of ascorbic acid. Additionally, the CeO2NPs showed 65.2% anti-inflammatory activity at the same concentration, compared to 70.1% for a standard drug. Antibacterial testing revealed that the CeO2NPs were particularly effective against multi-drug resistant strains, includingPseudomonas aeruginosa,Enterococcus faecalis, and MRSA, with moderate activity againstKlebsiella pneumoniae. These findings suggest that CeO2NPs synthesized viaT. terrestrishave strong potential as antimicrobial agents in addressing MDR infections.

IncC plasmid genome rearrangements influence the vertical and horizontal transmission tradeoff in Escherichia coli.

It has been shown that an evolutionary tradeoff between vertical (host growth rate) and horizontal (plasmid conjugation) transmissions contributes to global plasmid fitness. As conjugative IncC plasmids are important for the spread of multidrug resistance (MDR), in a broad range of bacterial hosts, we investigated vertical and horizontal transmissions of two multidrug-resistant IncC plasmids according to their backbones and MDR-region rearrangements, upon plasmid entry into a new host. We observed plasmid genome deletions after conjugation in three diverse natural Escherichia coli clinical strains, varying from null to high number depending on the plasmid, all occurring in the MDR region. The plasmid burden on bacterial fitness depended more on the strain background than on the structure of the MDR region, with deletions appearing to have no impact. Besides, we observed an increase in plasmid transfer rate, from ancestral host to new clinical recipient strains, when the IncC plasmid was rearranged. Finally, using a second set of conjugation experiments, we investigated the evolutionary tradeoff of the IncC plasmid during the critical period of plasmid establishment in E. coli K-12, by correlating the transfer rates of deleted or non-deleted IncC plasmids and their costs on the recipient strain. Plasmid deletions strongly improved conjugation efficiency with no negative growth effect. Our findings indicate that the flexibility of the MDR-region of the IncC plasmids can promote their dissemination, and provide diverse opportunities to capture new resistance genes. In a broader view, they suggest that the vertical-horizontal transmission tradeoff can be manipulated by the plasmid to improve its fitness.

Adjuvants restore colistin sensitivity in mouse models of highly colistin-resistant isolates, limiting bacterial proliferation and dissemination.

Antimicrobial resistance (AMR) has led to a marked reduction in the effectiveness of many antibiotics, representing a substantial and escalating concern for global health. Particularly alarming is resistance in Gram-negative bacteria due to the scarcity of therapeutic options for treating infections caused by these pathogens. This challenge is further compounded by the rising incidence of resistance to colistin, an antibiotic traditionally considered a last resort for the treatment of multi-drug resistant (MDR) Gram-negative bacterial infections. In this study, we demonstrate that adjuvants restore colistin sensitivity in vivo. We previously reported that the salicylanilide kinase inhibitor IMD-0354, which was originally developed to inhibit the human kinase IKKβ in the NFκB pathway, is a potent colistin adjuvant. Subsequent analog synthesis using an amide isostere approach led to the creation of a series of novel benzimidazole compounds with enhanced colistin adjuvant activity. Herein, we demonstrate that both IMD-0354 and a lead benzimidazole effectively restore colistin susceptibility in mouse models of highly colistin-resistant Klebsiella pneumoniae and Acinetobacter baumannii-induced peritonitis. These novel adjuvants show low toxicity in vivo, significantly reduce bacterial load, and prevent dissemination that could otherwise result in systemic infection.

Zidovudine in synergistic combination with nitrofurantoin or omadacycline: in vitro and in murine urinary tract or lung infection evaluation against multidrug-resistant Klebsiella pneumoniae.

Limited treatment options and multidrug-resistant (MDR) Klebsiella pneumoniae present a significant therapeutic challenge, underscoring the need for novel approaches. Drug repurposing is a promising tool for augmenting the activity of many antibiotics. This study aimed to identify novel synergistic drug combinations against K. pneumoniae based on drug repurposing. We used the clinically isolated GN 172867 MDR strain of K. pneumoniae to determine the reversal resistance activity of zidovudine (AZT). The combined effects of AZT and various antibiotics, including nitrofurantoin (NIT) and omadacycline (OMC), were examined using the checkerboard method, growth curves, and crystal violet assays to assess biofilms. An in vitro combination activity testing was carried out in 12 isolates of K. pneumoniae. In vivo murine urinary tract and lung infection models were used to evaluate the therapeutic effects of AZT + NIT and AZT + OMC, respectively. The fractional inhibitory concentration index and growth curve demonstrated that AZT synergized with NIT or OMC against K. pneumoniae strains. In addition, AZT + NIT inhibited biofilm formation and cleared mature biofilms. In vivo, compared with untreated GN 172867-infected mice, AZT + NIT and AZT + OMC treatment decreased colony counts in multiple tissues (P < 0.05) and pathological scores in the bladder and kidneys (P < 0.05) and increased the survival rate by 60% (P < 0.05). This study evaluated the combination of AZT and antibiotics to treat drug-resistant K. pneumoniae infections and found novel drug combinations for the treatment of acute urinary tract infections. These findings suggest that AZT may exert significant anti-resistance activity.

Molecular characterization and genotype of multi-drug resistant Staphylococcus epidermidis in nasal carriage of young population, Mahasarakham, Thailand.

Staphylococcus epidermidis, a coagulase-negative staphylococcus, is a prevalent skin commensal that has increasingly been recognized as a significant pathogen, particularly in hospital environments, where it is associated with device-related infections. The emergence of multi-drug resistance and its ability to form biofilms complicate the clinical management of infections caused by this organism, posing a growing public health concern. This study aimed to investigate the nasal carriage of S. epidermidis among healthy young individuals and to analyze its antibiotic resistance patterns, resistance genes, and biofilm formation capabilities. Nasal swabs were collected from 40 undergraduate students at Mahasarakham University, Thailand, aged between 20 and 22 years. A total of 38 isolates were confirmed as S. epidermidis through both phenotypic and molecular characterization. Antibiotic susceptibility testing demonstrated resistance to various classes of antimicrobials, including beta-lactams, macrolides, and tetracyclines. Notably, five isolates exhibited methicillin resistance S. epidermidis (MRSE). Resistance genes, such as mecA, ermA, tetM, tetL, and tetK, were identified across the isolates, contributing to the observed resistance profiles. Biofilm formation assays revealed that most isolates displayed weak to moderate biofilm formation, with only one isolate demonstrating strong biofilm-forming capacity. Genetic analysis indicated a significant correlation between biofilm formation and the presence of the icaA gene, which is crucial for biofilm production. This study suggests the necessity for ongoing surveillance of nasal carriage of S. epidermidis to enhance understanding of its role in the dissemination of antimicrobial resistance and biofilm-associated infections, particularly within healthcare settings.

Risk factors for multidrug-resistant bacteria in critically ill children and MDR score development.

Antimicrobial resistance and healthcare-associated infections (HAIs) are major health concerns in the pediatric intensive care unit (PICU). Device-associated HAIs (DA-HAIs) produced by multidrug-resistant (MDR) bacteria are especially worrying, as they can lead to an inappropriate empirical antibiotic therapy, worsened outcomes and increased mortality. The MDR score was designed to enable the prompt identification of patients at high risk of developing an MDR infection. This was a single-center, prospective, observational study, conducted between January 2015 and December 2022, including PICU patients with a microbiologically confirmed DA-HAI. Demographic, clinical characteristics and outcomes were compared between patients with a DA-HAI caused by MDR and non-MDR-associated DA-HAI, and a risk score for multi-resistance was designed. In total, 257 DA-HAI cases were included, 86 (33.46%) caused by an MDR microbe. In the univariate analysis, comorbidity (p = 0.002), previous MDR colonization (p < 0.001), previous surgery (p = 0.018), and previous antibiotic therapy (p = 0.009) were more frequent among MDR-associated DA-HAI (MDR DA-HAI). In addition, days from device insertion to infection and from PICU admission (p < 0.005) to infection were longer in patients with MDR. In the multivariate analysis, previous comorbidity (OR 2.201), previous MDR colonization (OR 5.149), and PICU length of stay longer than 9days (OR 1.782) were independently associated with MDR-DA-HAI. Using these three independent risk factors for MDR, a risk score was created: the MDR score. Three risk groups were obtained: low risk (0-2 points), intermediate risk (3-7 points), and high risk (8-12 points). Seventy-one patients with MDR-DA-HAI (82.6%) were classified in the intermediate or high-risk group, with a global sensitivity of 82.6%. The specificity in the high-risk group was 91.8%, and 81.0% of patients who were stratified into the low-risk group had non-MDR-associated infections, so they were correctly classified. Conclusions: The MDR score can be a useful tool to stratify patients in risk groups for MDR-DA-HAI. It may help to guide the choice of empirical therapy, leading to early optimization and avoiding delays in establishing appropriate treatment. This study reinforces the importance of stratifying patients based on their individual risk profile for MDR infection.

Synergistic invitro antimicrobial activity of polyherbal combination of Morinda lucida fruit and Pterocarpus santalinoides seed against multi-drug resistant clinical bacterial isolates

Background: Synergistic drug combination has been shown to be a way of bypassing drug resistance and reducing the amounts of antimicrobials consumed. As infections caused by multi-drug resistant organisms (MDROs) continue to pose global threat, it is important to search for new antimicrobial combinations of plant origin that are safe and readily available. The objective of this study is to evaluate the synergistic antimicrobial activity of extracts of Morinda lucida fruit and Pterocarpus santalinoides seed against multi-drug resistant (MDR) bacterial isolates Methodology: Morinda lucida and P. santalinoides fruits were plucked and washed clean, and the fruits of P. santalinoides were deseeded to remove the seeds. They were cut into smaller pieces and dried under shade before being milled into smooth powder and extracted with methanol. The clinical bacterial isolates used were MDR Escherichia coli, Proteus mirabilis, Klebsiella pneumoniae, Pseudomonas aeruginosa, and Staphylococcus aureus. The phytochemical constituents of the extracts were determined using the standard method. The invitro antimicrobial assay of the extracts was done at a concentration of 50 to 400 mg/ml using the agar well diffusion technique. The minimum inhibitory concentration (MIC) and fractional inhibitory concentration (FIC) of each extract were determined using the Checker-board micro-titration method, and the FIC result obtained for each isolate was used to calculate the fractional inhibitory concentration index (FICI) of the combined extracts. The time kill assay was performed to confirm the synergistic and bactericidal activities from the MIC values obtained. Results: The phytochemical analysis revealed that the extracts of both herbal plants contained alkaloids, phenol, tannin, saponin, glycosides, and terpenoids. The antimicrobial assay results showed that the polyherbal combination of the extracts was active against all the MDR bacterial isolates tested with varying zones of inhibition. The mean inhibition zone diameters of individual M. lucida and P. santalinoides ranged from 18.0±0.9mm to 26.0±1.4mm and 17.0±0.1mm to 24.0±0.2mm respectively, while the MIC ranged from 6.25mg/ml to 12.5mg/ml for M. lucida and 12.5 mg/ml for P. santalinoides. The mean inhibition zone diameter of the polyherbal combination of the two plant extracts ranged from 25.00±00mm to 34.0±0.4mm, which compared favorably with that of levofloxacin control (28.0±0.0mm to 32.00±0.0mm), while their MIC ranged from 0.39mg/ml to 1.56mg/ml. The FIC of M. lucida extract ranged from 0.06mg/ml to 0.25mg/ml while that of P. santalinoides ranged from 0.03mg/ml to 0.13mg/ml. The FICI of combined extracts ranged from 0.09mg/ml to 0.38 mg/ml for all the MDR isolates, which is less than 0.5mg/ml, indicating synergism against all the isolates. The time of kill assay confirmed the synergistic and bactericidal activities with a 3log10 CFU/ml decrease in the number of viable cells within 6 hours of incubation.Conclusion: Our findings showed synergistic antibacterial actions of extracts of M. lucida fruit and P. santalinoides seed against MDR clinical bacterial isolates, comparable to levofloxacin. Combination of these herbal plants may serve as alternative sources of antimicrobial agents for the treatment of infections caused by MDR bacterial pathogens.

Characterization of Chlorhexidine Resistance Among Clinical Isolates of Coagulase-Negative Staphylococcus Species in a Tertiary Care Center, Chennai.

Background Coagulase-negativeStaphylococci(CoNS) are potential pathogens and are often associated with healthcare-associated infections (HAIs). Chlorhexidine (CHX) is the most widely used antiseptic to reduce colonization and infection by allStaphylococci, including CoNS. Resistance to CHXamong CoNS has been observed over the past few years, consequent to its widespread use. Phenotypic tolerance or reduced susceptibility to CHX is conferred by plasmid-mediatedqacgroup of genes, mainlyqacA/Bandsmr, which cause activation of efflux pumps over the bacterial cell wall. This study aims to characterize the phenotypic and genotypic resistance exhibited by CoNS species against CHX. Methods After ethical approval, 148 consecutive, non-repetitive isolates of clinically significant CoNS species of hospitalized patients, isolated from blood samples and exudative specimens, were included in the study. Speciation was performed by conventional biochemical identification and automated methods. Antimicrobial susceptibility testing was performed by disc diffusion technique and for vancomycin by minimum inhibitory concentration (MIC) determination, as per Clinical Laboratory Standards Institute (CLSI) M-100 2023 guidelines. Methicillin resistance was detected using a cefoxitin disc. MIC for CHX was performed by agar dilution method; reduced susceptibility was considered when MIC to CHX ≥4 µg/mL. The simplex polymerase chain reaction (PCR) was carried out with suitable controls to detectqacA/Bandsmr. Statistical analysis was conducted to determine the association ofqacA/Bandsmrgenes with MIC of CHXin the study isolates. Results Fifteen different species of CoNS were obtained from clinical samples. A high percentage of resistance was observed against various classes of antibiotics. Methicillin resistance was observed in 69.6% (103/148) of isolates. Of 148 CoNS, 52.7% (78/148) of isolates exhibited reduced susceptibility to CHX with an MIC ≥4 µg/mL. These isolates exhibited a higher percentage of methicillin resistance (75.6%, 59/78). By PCR, 34.5% (51/148) of isolates carried either or both genes. GeneqacA/Bwas solely detected in 27.02% (40/148) of isolates, of which 14 were CHX-tolerant and the remaining 26 were CHX-susceptible. Genesmrwas solely detected in 4.1% (6/148) of isolates comprising three isolates each in CHX-tolerant and susceptible categories.There were 3.4% (5/148) of isolates that harbored bothgenes, of which only one isolate was CHX-susceptible, while the other four were CHX-tolerant. A proportion of isolates that were phenotypically tolerant to CHX did not carry either or both genes. A significant statistical association was found between reduced susceptibility to CHX and the presence of antiseptic resistance genesin the study isolates (p-value=0.033942). Conclusion To our knowledge, this is the first study from South India to investigate CHX resistance among CoNS using phenotypic and genotypic methods. The rise of antiseptic resistance among CoNS is an emerging threat to current infection control practices. The presence ofqacA/Bandsmrgenes, especially in CHX susceptible isolates, is concerning since these resistance genes are located on transferable plasmids, and the isolates can develop resistance eventually upon exposure to CHX.

Phage-liposome nanoconjugates for orthopedic biofilm eradication

Infection by multidrug-resistant (MDR) bacteria has become one of the biggest threats to public health worldwide. One reason for the difficulty in treatment is the lack of proper delivery strategies into MDR bacterial biofilms, where the thick extracellular polymeric substance (EPS) layer impedes the penetration of antibiotics and nanoparticles. Here, we propose a novel bioactive nanoconjugate of drug-loaded liposomes and bacteriophages for targeted eradication of the MDR biofilms in orthopedic infections. Phage Sb-1, which has the ability to degrade EPS, was conjugated with antibiotic-loaded liposomes. Upon encountering the biofilm, phage Sb-1 degrades the EPS structure, thereby increasing the sensitivity of bacteria to antibiotics and allowing the antibiotics to penetrate deeply into the biofilm. As a result, effective removal of MDR bacterial biofilm was achieved with low dose of antibiotics, which was proved in this study by both in vitro and in vivo investigations. Notably, in the rat prosthetic joint infection (PJI) model, we found that the liposome-phage nanoconjugates could effectively decrease the bacterial load in the infected area and significantly promote osteomyelitis recovery. It is therefore believed that the conjugation of bacteriophage and liposomes could open new possibilities for the treatment of orthopedic infections, possibly other infections in the deep tissues.

Safety assessment of enterocin-producing Enterococcus strains isolated from sheep and goat colostrum

BackgroundThis study investigates the safety evaluation of enterocin-producing 11 E. mundtii and two E. faecium strains previously isolated from small livestock colostrums. Enterococcus species do not possess Generally Recognized as Safe (GRAS) status. Hence, it is critical to scrutinize enterococci’s antibiotic resistance, virulence characteristics, and biogenic amine production capabilities in order to assess their safety before using them as starter or adjunct cultures.ResultsEnterococcus strains showed susceptibility to medically significant antibiotics. Multiple-drug resistance (MDR) was found in only E. faecium HC121.4, and its multiple antibiotic resistance (MAR) index was detected to be 0.22. The tetL and aph(3')-IIIa were the most commonly found antibiotic resistance genes in the strains. However, E. mundtii strains HC56.3, HC73.1, HC147.1, and E. faecium strain HC121.4 were detected to lack any of the antibiotic resistance genes examined in this study. Only E. mundtii HC166.3 showed hemolytic activity, while none of the strains engage in gelatinase activity. The strains were identified to have virulence factor genes with a low rate. None of the virulence factor genes could be detected in E. mundtii HC26.1, HC56.3, HC73.1, HC165.3, HC166.8, and E. faecium HC121.4. The E. mundtii HC73.2 strain displayed the highest presence of virulence factor genes, namely gelE, efaAfs, cpd, and ccf. Similarly, the E. mundtii HC112.1 strain showed a significant presence of genes efaAfm, ccf, and acm. There was no decarboxylation of histidine, ornithine, or lysine seen in any of the strains. Nevertheless, E. faecium HC121.4 and HC161.1 strains could decarboxylate tyrosine, but E. mundtii HC26.1, HC56.3, HC73.1, HC73.2, HC112.1, HC147.1, HC155.2, HC165.3, HC166.3, HC166.5, and HC166.8 strains only showed a limited capacity for tyrosine decarboxylation. None of the strains possessed the hdc, odc, or ldc genes, but all of them had the tdc gene.ConclusionThe E. mundtii HC56.3 and HC73.1 strains were deemed appropriate for utilization in food production. Using the remaining 11 strains as live cultures in food production activities could pose a possible risk to consumer health.

In vitro evaluation of Morinda lucida root extracts against multi-drug resistant bacterial pathogens isolated from diabetic foot ulcers

Background: The continuous rise in microbial resistance to orthodox antimicrobial drugs has led to the search for alternative sources with proven efficacy to solve the challenges of antimicrobial resistance (AMR). The preferred alternatives are plant sources, and this has led to the evaluation of constituents and potency of medicinal plants to provide scientific justification for their use. Methodology: The root of Morinda lucida plant was dug up from the ground, washed clean, and cut into smaller pieces and dried. The root was then ground into fine powder and extracted with water (aqueous), methanol, ethyl acetate and n-hexane. Multi-drug resistant (MDR) Staphylococcus aureus, Escherichia coli, Klebsiella pneumoniae, Proteus mirabilis and Pseudomonas aeruginosa were isolated from wound swab samples collected from patients with diabetic foot ulcers using conventional cultures, biochemical identification, and antimicrobial susceptibility tests. The phytochemical and proximate contents of the extracts were assessed by standard technique. The in vitro antimicrobial activities of the extracts were determined at a concentration of 200mg/ml of the extract using the agar well diffusion technique. The minimum inhibitory and bactericidal concentrations were determined by serial doubling dilution. The methanol extract time-kill assay was performed to determine the time of kill of the bactericidal concentration. Results: The phytochemical analysis showed that M. lucida root contains essential secondary metabolites such as flavonoids, alkaloids, tannin, saponin, glucosides, anthraquinone and quinine. The methanol and aqueous extracts showed higher in vitro antibacterial activity, producing the highest zone of inhibition of 27mm against S. aureus but a lower activity of 18mm with n-hexane extract against all isolates except S. aureus. The MIC ranges from 3.125mg/ ml and 25mg/ml. The time-kill assay of methanol extract at 2x and 3x MIC showed that bactericidal activity occurred within 0-8 hours of incubation, indicating high activity. Conclusion: The antibacterial potency of M. lucida root extract and the phytochemical components from this study shows that it can serve as a source of alternative antimicrobial agent that may be effective in the treatment MDR bacterial infections.

Genetic relatedness of diarrheagenic Escherichia coli pathotypes isolated from children under five years of age and food animals in Kisumu County, Kenya

Background: Diarrheal disease remains one of the leading causes of deaths in children below five years of age. The risk factors associated with diarrhea include poor hygiene practices such as open defecation and consumption of contaminated water and food. However, exposure of domestic animal is equally a potential risk factor for diarrhea disease in children. Methodology: We characterized animal-related exposures in a subset of households (n=73) by collecting faecal samples from 150 children with diarrhoea and 100 food animals (30 cattle, 30 chicken, 25 goats and 15 pigs). Escherichia coli was isolated from the faecal samples and biochemically confirmed using conventional microbiological techniques. The deoxyribonucleic acid (DNA) of each E. coli isolate was extracted and amplified by multiplex PCR to identify three diarrheagenic E. coli pathotypes. The amplified products were sequenced, and genetic relatedness of the isolates was determined through phylogenetic analysis. Results: We isolated and identified a total of 32 (12.8%) diarrheagenic E. coli (DEC) from the 250 faecal samples, 26 (17.3%) of which were from the 150 children with diarrhea while 6 (6.0%) were from the 100 food animals (OR=3.285, 95% CI=1.299-8.305, p=0.011). Three DEC pathotypes were confirmed by PCR in 16 DEC strains, with 9 enteroaggregative E. coli (EAEC), 2 enterotoxigenic E. coli (ETEC), 2 enteropathogenic E. coli (EPEC), 1 EAEC/ETEC, 1 EAEC/EPEC and 1 ETEC/EPEC mixed strains. The phylogenetic analysis showed that 6 DEC isolates had genetic similarity ranging between 31% to 90%. Isolates S04 originating from animal and S02 from a child with diarrhoea of the same household were closely related, with 55% similarity. Moreover, isolate S05 from animal origin and S06 of diarrheic child origin were closely related, with similarity degree as high as 82% even though they were not paired. Twenty four of the 26 (92.3%) DEC isolates from diarrhoeic children showed multidrug resistance (MDR) pattern to antibiotics but none of the 6 isolates from food animals was multi-drug resistant. Conclusion: The high degree of genetic relationship between DEC isolate S04 and S02 from animal and human origin indicated the high potency of zoonotic transmission. Further studies investigating animal husbandry practices and zoonotic transmission of DEC are needed.

Myricetin inhibits CYP3A4, GST, and MRP1 in hepatic cancer cells

AbstractHerbal and nutritional supplements are widely used to prevent and treat many diseases, including cancer. Tumor cells modify metabolic enzyme systems like CYP3A4 and GST. They also overexpress MRP1, an ATP-binding cassette transporter subfamily G (ABCG2) member. Drug efflux may increase, reducing tumor cell drug accumulation and developing drug resistance that leads to significant obstacles in cancer care. Natural products' ability to overcome cancer's multidrug resistance is interesting. Their ability to affect several targets makes them valuable in addressing drug resistance from diverse approaches. The potential of natural flavonoid; Myricetin (MYR) to modulate CYP3A4, GST, and MRP1 activity and expression in hepatic cancer cells was evaluated to prove its targeting and preventing these pathways of multidrug resistance. The cell proliferation of MYR was determined using an MTT assay. Specific enzyme assays, efflux assay, and gene expression using RT-PCR were used to evaluate MYR effect in hepatic cell lines HepG-2 and Huh-7. MYR has a noteworthy cytotoxic effect compared to doxorubicin (DOX) with IC50 &gt; 100 μM in HepG-2 and Huh-7 cells. MYR showed potent inhibition of CYP3A4 and GST enzyme activity and MRP1 efflux function and downregulated their gene expression in a dose-dependent manner in both cells. MYR100 dose was the most significant effective dose. MRY100 decreased CYP3A4 activity by 67.5% (P &lt; 0.05) and 55% (P &lt; 0.01) and downregulated the gene by 0.2-fold (P &lt; 0.001) and 0.3-fold (P &lt; 0.001) in HepG-2 and Hub-7 cells, respectively. After treatment with MRY100, GST activity decreased significantly in both cells, reaching 47.6% (P &lt; 0.001) and 33.2% and GST gene downregulation was 0.12 and 0.21-fold (P &lt; 0.001). MRY100 inhibited MRP1 efflux pump 2.3 times (P &lt; 0.001) and 1.9 times (P &lt; 0.001) more effectively than PC, resulting in a 0.23-fold and 0.12-fold downregulation of MRP1 genes in HepG-2 and Hub-7 cells. The result will validate the use of MYR to interact with the metabolism phases and could be used as adjuvant therapy in cancer prevention and treatment approaches.

Disinfectants efficacy in reducing pathogens related to health-care infection associated in universities hospitals of Gorgan, North of Iran

BackgroundDisinfection has a fundamental role in the control of pathogens in the hospital environment. This study was designed to assess the efficacy and functional impact of disinfectants in reducing pathogens related to healthcare associated infections (HAIs) in hospitals.MethodsThis observation study was conducted at three university hospitals in Gorgan, Iran, from May to Oct 2023. The data including used disinfectants and microbiological examination were obtained from the infection control unit of each hospital.ResultsThe results showed that a variety of disinfectants from intermediate to high levels were employed in accordance with the World Health Organization (WHO) protocols. The microbial result revealed that 31.6% (286 out of 906) of the sample had at least one microorganism. Among identified organisms, Bacillus spp. were the predominant species followed by Staphylococcus epidermis, fungus genera, Enterobacter spp., Enterococcus spp., Pseudomonas spp., Escherichia coli, Alcaligenes spp., Staphylococcus aureus, Citrobacter spp., Corynebacterium spp., Klebsiella spp., Acinetobacter spp., Micrococcus spp., Staphylococcus saprophyticus, and Serratias spp. The highest prevalence rates of microorganisms were observed in the wards of ICU, emergency, internal medicine, and women’s ward. The chi-square test revealed a significant relationship between the presence of organisms and hospital wards (P < 0.05).ConclusionThe presence of pathogens indicates a defect in the disinfection process, probably due to both little attention to disinfection protocols and multidrug resistance. It is not yet possible to eliminate pathogens from the hospital environment, but it can be minimized by education intervention, standardizing disinfecting processes, and monitoring by the infection control committee.

Eco-friendly Biosynthesis of Silver Nanoparticles Using Elephantorrhiza elephantina: Characterization and Antimicrobial Activity against Mycobacterium tuberculosis Surrogate Models

The emergence of multidrug-resistant (MDR) and extensively-drug resistant (XDR) Mycobacterium tuberculosis (MTB) strains have scampered efforts to arrest the tuberculosis (TB) pandemic due to treatment failures based on current drugs. The need for novel therapeutics has never been more urgent. Research is leaning towards alternative composite antimicrobials such as silver nanoparticles (AgNPs). Elephantorrhiza elephantina (E. elephantina) is an indigenous Southern African rhizomatous plant which has been used for eons in African traditional medicine for the management of TB. The purpose of this study was to biosynthesize and characterize AgNps using E. elephantina, and to evaluate their antimicrobial effectiveness against surrogate MTB species. The study also sought to screen for the participating secondary metabolites in E. elephantina acting as bio reducing, capping and stabilising agents; and to determine the toxicity of the lyophilised root extract in vivo. Qualitative phytochemical screening techniques confirmed the presence of bioactive secondary metabolites with functional groups capable of mediating in the biosynthesis of AgNPs. UV-Vis spectrometry established the identity of the nanostructures as AgNPs. Transmission electron microscopy and dynamic light scattering confirmed the AgNPs to have an average size of 44 nm, with spherical and cubic morphologies. The biosynthesised AgNPs were potent inhibitors of the model MTB species with an MIC of 19.53 µg/mL, and inhibition zones of 20mm and 18mm against Mycolicibacterium aurum and Mycolicibacterium smegmatis respectively at their MICs. Oral toxicity evaluations based on OECD guideline 425 determined the LD50 of the lyophilised E. elephantina root extract to be safe above 4000mg/kg body weight. The study concluded that biosynthesized AgNPs from E. elephantina root extract, using a one-pot green-synthesis technique, were both safe and effective, offering a promising alternative therapeutic strategy for combating multidrug-resistant TB.

Bedaquiline versus injectable containing regimens for rifampicin-resistant and multidrug-resistant tuberculosis in a reference center in Brazil – a real-world evidence study using a retrospective design

BackgroundDrug resistance (DR) is one of the several challenges to global tuberculosis (TB) control. The implementation of bedaquiline (BED) for DR-TB after more than 40 years was expected to improve treatment outcomes as well as microbiologic conversion and adverse events (AE) occurrence.MethodsRetrospective cohort study based on secondary data of patients with rifampicin-resistant (RR) or multidrug-resistant (MDR) TB reported to the Outpatient Clinic of Mycobacterial Diseases of the Thorax Diseases Institute – Federal University of Rio de Janeiro - Brazil, between 2016 and 2023. We aimed to evaluate microbiologic conversion, AE and TB treatment outcomes and compare them according to the treatment regimen used for RR/MDR-TB patients under routine conditions [Injectable Containing Regimens (ICR) versus BED Containing Regimens (BCR)]. Logistic regression and survival analysis using Cox regression and Kaplan Meier curve were used for statistical analysis.ResultsOf the 463 DR-TB patients notified during the study period, 297 (64.1%) were included for analysis (ICR = 197 and BCR = 100). Overall AEs were more frequent (83.7 vs. 16.3%, p < 0.001) and occurred earlier in the ICR group (15 days vs. 65 days, p = 0.003). There were no cases of cardiotoxicity requiring interruption of BED treatment. None of the regimens of treatment tested were associated with smear or culture conversion on Cox regression analysis (p = 0.60 and 0.88, respectively). BED-containing regimens were also associated with favorable outcomes in multivariable logistic regression [adjusted odds ratio (aOR) = 2.63, 95% confidence interval (CI)1.36–5.07, p = 0.004], as higher years of schooling, primary drug resistance, and no previous TB treatment. In the survival analysis, BCR was inversely associated with the occurrence of AE during treatment follow-up (aHR 0.24, 95% CI 0.14–0.41, p < 0.001). In addition, TB treatment regimens with BED were also associated with favorable outcomes (aHR 2.41, 95% CI 1.62–3.57, p < 0.001), along with no illicit drug use and primary drug resistance.ConclusionsThe implementation of a fully oral treatment for RR/MDR-TB in a reference center in Brazil was safe and associated with favorable outcomes under routine conditions, despite social, demographic, and behavioral factors that may influence TB treatment completion.

A global genome dataset for Salmonella Gallinarum recovered between 1920 and 2024

Salmonella enterica serovar Gallinarum (S. Gallinarum) is an avian-specific pathogen responsible for fowl typhoid, a severe systemic disease with high mortality in chickens. This disease poses a substantial burden to the poultry industry, particularly in developing countries like China. However, comprehensive genome datasets on S. Gallinarum are lacking. Here, we present the most extensive S. Gallinarum genome dataset, comprising 574 well-collated samples. This dataset consists of 366 genomes sequenced in our laboratory and 208 publicly available genomes, collected from various continents over the past century. Using in silico prediction, we categorized S. Gallinarum into three distinct biovars. Regarding antimicrobial resistance, 238 strains (41.5%) carried antimicrobial resistance genes (ARGs) with a total of 635 records, while 232 strains (40.4%) exhibited multi-drug resistance. Mobile genomic elements (MGEs) serve as critical drivers for ARGs. Our dataset includes 5,636 MGEs records, with most MGEs belonging to prophages and plasmids. This dataset expands our understanding of the genomic characteristics of S. Gallinarum, providing valuable resources for future genomic studies to improve disease management.

RpoS activates formation of Salmonella Typhi biofilms and drives persistence in the gall bladder.

The development of strategies for targeting the asymptomatic carriage of Salmonella Typhi in chronic typhoid patients has suffered owing to our basic lack of understanding of the molecular mechanisms that enable the formation of S. Typhi biofilms. Traditionally, studies have relied on cholesterol-attached biofilms formed by a closely related serovar, Typhimurium, to mimic multicellular Typhi communities formed on human gallstones. In long-term infections, S. Typhi adopts the biofilm lifestyle to persist in vivo and survive in the carrier state, ultimately leading to the spread of infections via the fecal-oral route of transmission. In the present work, we studied S. Typhi biofilms directly, applied targeted as well as genome-wide genetic approaches to uncover unique biofilm components that do not conform to the CsgD-dependent pathway established in S. Typhimurium. We undertook a genome-wide Tn5 mutation screen in H58, a clinically relevant multidrug resistance strain of S. Typhi, in gallstone-mimicking conditions. We generated New Generation Sequencing libraries based on the ClickSeq technology to identify the key regulators, IraP and RpoS, and the matrix components Sth fimbriae, Vi capsule and lipopolysaccharide. We discovered that the starvation sigma factor, RpoS, was required for the transcriptional activation of matrix-encoding genes in vitro, and for S. Typhi colonization in persistent infections in vivo, using a heterologous fish larval model. An rpoS null mutant failed to colonize the gall bladder in chronic zebrafish infections. Overall, our work uncovered a novel RpoS-driven, CsgD-independent paradigm for the formation of cholesterol-attached Typhi biofilms, and emphasized the role(s) of stress signaling pathways for adaptation in chronic infections. Our identification of the biofilm regulators in S. Typhi paves the way for the development of drugs against typhoid carriage, which will ultimately control the increased incidence of gall bladder cancer in typhoid carriers.