non-MDR Isolates Research Articles

Detection of Efflux pump MexX and MexY genes in Multidrug Resistant Pseudomonas aeruginosa

High antibiotic resistance among Pseudomonas aeruginosa could be viewed as the primary cause of nosocomial infections. In this study, 60 isolates of Pseudomonas aeruginosa were isolated from urine, sputum, burn, ear and wounds swab. The major goal was to determine the relationship between MexX and MexY and the resistance of MDR P. aeruginosa to many drugs. Prevalence of multidrug resistant (MDR) Pseudomonas aeruginosa was 22 isolates (36.66%) and 38 isolates (63.34%) were found to be non-MDR isolates. Moreover, molecular detection of MexX and MexY genes was done for MDR isolates.MexY gene was found in all MDR isolates and MexX gene was detected in 20 (90.9%) MDR isolates. Susceptibility test showed highly significant differences p≤0.01. Highest resistance was found against piperacillin/sulbactam (45%), followed by ceftazidime/avibactam with 33.33%, and by ceftolozane/tazobactam and levofloxacin which shared same percentage at 31.67%. Almost 23.33% were less resistant to both doripenem and colistin, and 25% to tobramycin. All these findings confirmed a positive correlation between the presence of MexX and MexY genes and an increase in the resistance for all antibiotics used in this study the results of which were highly significant at p≤0.01 for all antibiotics. The study found that some antibiotics that had not been studied previously, also had a relationship with the presence of these genes and the resistance of bacteria to antibiotics such as penicillin, unlike it was previously believed that they do not have relation with the efflux pump mechanism in P. aeruginosa and that this bacterium use other mechanisms to resist them.

Emergence of extensively drug-resistant hypervirulent Acinetobacter baumannii isolated from patients with bacteremia: bacterial phenotype and virulence analysis

ObjectivesIndividuals infected with extensively drug-resistant (XDR) Acinetobacter baumannii are difficult to cure and have a high mortality rate. In this study, we compared the genomic and phenotypic differences between XDR and non-multidrug-resistant (MDR) A. baumannii and further characterized hypervirulent XDR A. baumannii. MethodsA total of 1,403 Acinetobacter isolates were collected from patients with bacteremia between 1997 and 2015. Antimicrobial susceptibility test was performed to categorize isolates into non-MDR, MDR, and XDR groups. The presence of selected virulence-associated genes was determined by PCR. Bacterial phenotypes, including iron acquisition, biofilm formation, capsule production, and virulence to larvae and mice, were determined. ResultsMultilocus sequence types revealed the high prevalence of ST2 (81.6%) and ST129 (18.4%) among 49 XDR isolates and sequence types of 18 non-MDR isolates were more diverse. Virulence-associated phenotypic assays showed that XDR isolates had higher iron acquisition ability and capsule production and higher virulence to Galleria mellonella larvae. However, their biofilm formation ability was lower compared to that of non-MDR isolates. XDR isolates contained more virulence genes, tonB, hemO, abaI, and ptk, while non-MDR isolates contained more pld and ompA. Twenty-one XDR isolates caused <20% larvae survival rate after 7 days post-infection were defined as hypervirulent XDR isolates. Among them, isolates 1677 (ST129) and 929-1 (ST2) also caused the death of all infected mice within two days. ConclusionSome subpopulations of highly drug-resistant ST2 also exhibit high virulence. Therefore, it is of utmost importance to continue monitoring the spread of hypervirulent XDR A. baumannii isolates.

Assessment of biofilm-forming capacity and multidrug resistance in Staphylococcus aureus isolates from animal-source foods: implications for lactic acid bacteria intervention

BackgroundStaphylococcus aureus, a Gram-positive bacterium, poses a significant threat to public health and food safety due to its virulence and its ability to develop antimicrobial resistance (AMR). Moreover, S. aureus can form biofilms in food environments, making it difficult to eradicate and pose a major challenge in foodborne illness prevention.MethodsThe study aimed to investigate the biofilm-forming capabilities and AMR profiles of 107 S. aureus isolates derived from milk, chicken meat, and chicken eggs. Further, the study compared the biofilm formation tendencies between multi-drug resistant (MDR) and non-MDR S. aureus isolates. Additionally, the research explored the antibacterial and anti-biofilm properties of Lactobacillus rhamnosus and Lactobacillus casei, focusing on their aggregation and co-aggregation effects with S. aureus.ResultsAround 70.10% of S. aureus isolates were found to be resistant to at least three antibiotic classes. The biofilm assay revealed that 16.82% isolates were strong biofilm formers. The MDR isolates displayed a strong biofilm-forming ability (i.e., 18.67%) and a higher prevalence of biofilm-associated genes [i.e., icaA (53.33%) and icaD (44.0%)] compared to non-MDR isolates. The LAB strain, L. rhamnosus exhibited a 29.06 mm mean antibacterial inhibition zone, an average reduction of 48.19% in biofilm growth, 55.46% auto-aggregation, and 40.61% co-aggregation with S. aureus. Similarly, L. casei demonstrated a 21.80 mm mean antibacterial inhibition zone, an average reduction of 31.56% in biofilm growth, 45.23% auto-aggregation, and 36.81% co-aggregation with S. aureus isolates.ConclusionThis study provides valuable insights into the biofilm formation of MDR S. aureus and underscores the potential of L. rhamnosus and L. casei as bio-control agents. These findings highlight the necessity for additional research into the mechanisms through which LAB strains inhibit pathogenic biofilms and their potential applications in enhancing food safety.

Characterization of antibiotic and disinfectant susceptibility in biofilm-forming Acinetobacter baumannii: A focus on environmental isolates.

The clinical role of Acinetobacter baumannii has been highlighted in numerous infectious syndromes with a high mortality rate, due to the high prevalence of multidrug-resistant (MDR) isolates. The treatment and eradication of this pathogen is hindered by biofilm-formation, providing protection from noxious environmental factors and antimicrobials. The aim of this study was to assess the antibiotic susceptibility, antiseptic susceptibility and biofilm-forming capacity using phenotypic methods in environmental A.baumannii isolates. One hundred and fourteen (n = 114) isolates were collected, originating from various environmental sources and geographical regions. Antimicrobial susceptibility testing was carried out using the disk diffusion method, while antiseptic susceptibility was performed using the agar dilution method. Determination of biofilm-forming capacity was carried out using a microtiter-plate based method. Resistance in environmental A. baumannii isolates were highest for ciprofloxacin (64.03%, n = 73), levofloxacin (62.18%, n = 71) and trimethoprim-sulfamethoxazole (61.40%, n = 70), while lowest for colistin (1.75%, n= 2). Efflux pump overexpression was seen in 48.25% of isolates (n = 55), 49.12% (n = 56) were classified as MDR. 6.14% (n = 7), 9.65% (n = 11), 24.65% (n = 28) and 59.65% (n = 68) of isolates were non-biofilm producers, weak, medium, and strong biofilm producers, respectively. No significant differences were observed between non-MDR vs. MDR isolates regarding their distribution of biofilm-producers (P = 0.655). The MIC ranges for the tested antiseptics were as follows: benzalkonium chloride 16-128 μg mL-1, chlorhexidine digluconate 4-128 μg mL-1, formaldehyde 64-256 μg mL-1 and triclosan 2-16 μg mL-1, respectively. The conscientious use of antiseptics, together with periodic surveillance, is essential to curb the spread of these bacteria, and to maintain current infection prevention capabilities.

Assessment of Antibiotic Resistance among Clinical Isolates of Enterobacteriaceae in Nepal.

Clinicians face a global challenge treating infections caused by Enterobacteriaceae because of the high rate of antibiotic resistance. This cross-sectional study from the Nepal Armed Police Force Hospital, Kathmandu, Nepal, characterized resistance patterns in Enterobacteriaceae across different antimicrobial classes and assessed incidences of multidrug-resistant (MDR) and extensively drug-resistant (XDR) infections. Enterobacteriaceae from clinical samples were isolated on blood and MacConkey agar, except for urine samples on cysteine lactose electrolyte-deficient agar. To determine antimicrobial susceptibility patterns, including MDR and XDR, the Kirby-Bauer disc diffusion method was used. Statistics were performed using SPSS, v. 17.0. Members of the family were identified in 14.5% (95% CI: 16.2-12.8%) of the total samples (N = 1,617), primarily in urine (54.7%, 128/234), blood (19.7%, 46/234), and sputum (15.0%, 35/234). Escherichia coli (n = 118, 44.2%) was the most predominant bacteria, followed by Citrobacter freundii (n = 81, 30.3%). As much as 95.6% (392/410) of the isolates were penicillin-resistant, whereas only 36.2% (290/801) were carbapenem-resistant. A total of 96 (36.0%) MDR and 98 (36.7%) XDR Enterobacteriaceae were identified. Proteus mirabilis (44.4%, 8/18) predominated MDR cultures, whereas C. freundii (53.1%, 43/81) predominated XDR cultures. Multidrug resistant (38.4%, 71/154) and XDR Enterobacteriaceae (22.7%, 35/154) were chiefly uropathogens. Fluoroquinolone resistance rates in non-MDR, MDR, and XDR isolates were 19.9%, 63.2%, and 96.2%, respectively, whereas cephalosporin resistance rates were 28.6%, 72.9%, and 95.4% and penicillin resistance rates were 67.0%, 97.4%, and 98.0%. One-seventh of patients visiting the hospital were found to be infected with Enterobacteriaceae, and of these patients, at least one-fourth were infected with MDR strains.

Comparative molecular profiling of multidrug-resistant Pseudomonas aeruginosa identifies novel mutations in regional clinical isolates from South India.

We sought to analyse the antibiotic susceptibility profiles and molecular epidemiology of MDR clinical Pseudomonas aeruginosa isolates from South India using non-MDR isolates as a reference. We established a comprehensive clinical strain library consisting of 58 isolates collected from patients across the South Indian state of Kerala from March 2017 to July 2019. The strains were subject to antibiotic susceptibility testing, modified carbapenem inactivation method assay for carbapenemase production, PCR sequencing, comparative sequence analysis and quantitative PCR of MDR determinants associated with antibiotic efflux pump systems, fluoroquinolone resistance and carbapenem resistance. We performed in silico modelling of MDR-specific SNPs. Of our collection of South Indian P. aeruginosa clinical isolates, 74.1% were MDR and 55.8% were resistant to the entire panel of antibiotics tested. All MDR isolates were resistant to levofloxacin and 93% were resistant to meropenem. We identified seven distinct, MDR-specific mutations in nalD, three of which are novel. mexA was significantly overexpressed in strains that were resistant to the entire test antibiotic panel while gyrA and gyrB were overexpressed in MDR isolates. Mutations in fluoroquinolone determinants were significantly associated with MDR phenotype and a novel GyrA Y100C substitution was observed. Carbapenem resistance in MDR isolates was associated with loss-of-function mutations in oprD and high prevalence of NDM (blaNDM-1) within our sample. This study provides insight into MDR mechanisms adopted by P. aeruginosa clinical isolates, which may guide the potential development of therapeutic regimens to improve clinical outcomes.

2180. Pseudomonas aeruginosa epidemic high-risk clones and their association with multidrug-resistant

Abstract Background Pseudomonas aeruginosa (PAE) is an opportunistic pathogen associated with a variety of hospital infections. Treatment of infections caused by PAE is challenging due to the emergence of multidrug-resistant (MDR), which is associated with increased morbidity, mortality, and healthcare costs. Clonal spread and the presence of resistance genes probably explain the successful propagation of MDR. In Ecuador, data on the molecular epidemiology, as well as circulating clones, are limited. Therefore, the objective of this study is to know the population structure of PAE by identifying clones in clinical samples in Quito-Ecuador. Methods A significant set (45) of randomly selected clinical PAE isolates, including multidrug and non-multidrug resistant isolates, were assigned to sequence types (STs) and compared them with their antibiotic susceptibility profile. The genetic diversity was assessed by applying the multilocus sequence typing (MLST) scheme (Figure 1) and the genetic relationships between different STs were corroborated by the phylogenetic neighbor-net network (Figure 2).Figure 1.Analysis of the allelic profiles using eBURST algorithm.Figure 2.Splits Tree showing the distribution of all the sequence types obtained for the clinical Pseudomonas aeruginosa isolates studied. The Splits Tree was based on the analysis of the allelic profiles of the acsA, aroE, guaA, mutL, nuoD, ppsA and trpE genes. Asterisk mark (*) indicates the MDR isolates Results The analysis of seven loci, acsA, aroE, guaA, mutL, nuoD, ppsA and trpE, identified 24 different ST and the most prevalent STs were ST-3750 and ST-253. A SplitsTree was constructed with all the isolates analyzed. The majority of the MDR isolates were included in ST-3750 and ST-253, also 3 singleton ST was identified in to MDR isolates. The 20 different ST was found to non-MDR isolates, and only 3 ST were found in more the one isolates. Conclusion The population structure of clinical PAE presents in these isolates indicate a significant association between MDR isolates and the clonal types: all ST-3750 and ST-253 isolates were MDR. ST-3750 is closely related strains to the ST111 (CC111), and ST-253 and ST111 are a group of successful high-risk clones and widely distributed worldwide. The multiresistant-isolates studied are grouped in the most prevalent sequence types found, and the susceptible isolates correspond mainly to singleton sequence types. Therefore, these high-risk clones and their association with multidrug resistance phenotypes are contributing to the spread of MDR in Quito-Ecuador. Disclosures Jeannete Zurita, n/a, Pfizer: Grant/Research Support

Prevalence of Type III Secretion System (T3SS) and Biofilm Development in Genetically Heterogeneous Clinical Isolates of Pseudomonas aeruginosa from Nigeria.

Pseudomonas aeruginosa infection in seriously ill patients is a major concern due to its ability to form biofilm and secrete effector toxins. There is little information on the prevalence of T3SS effector toxins and biofilm production in clinical isolates of P. aeruginosa from Nigeria. The goal of this study is to evaluate the prevalence of T3SS toxins and biofilm production among isolates from selected tertiary hospitals in Nigeria. This study examined 430 clinical isolates from our previous work, comprising 181 MDR (multidrug-resistant) and 249 non-MDR isolates. Biofilm production and type III secretion toxins were determined using colorimetric microtiter plate assay and polymerase chain reaction, respectively. Carbapenem-resistant isolates were typed using REP-PCR and BOX-PCR. Biofilm production was detected in 386/430 (89.8%) of the isolates. Out of 386 biofilm producers, 167 (43.3%) were multidrug-resistant isolates. PCR identified four T3SS virulence types among 430 isolates, including 78 (18.1%) exoU+/exoS- isolates, 343 (79.8%) exoU-/exoS + isolates, 5 (1.2%) exoU+/exoS + isolates, and 4 (0.9%) exoU-/exoS- isolates. Both REP- and BOX-PCR consist of eight clusters. On the REP-PCR dendrogram, ExoU+/ExoS- isolates majorly occupied cluster IV. Clusters IV, VII, and VIII consist of isolates from wounds on BOX-PCR dendrogram. There was a positive association between strong biofilm production and multidrug resistance in our P. aeruginosa isolates. This study identified multidrug-resistant, biofilm-producing P. aeruginosa strains that secrete cytotoxic effectors which are significant virulence factors in P. aeruginosa. This poses a severe risk to our healthcare system and highlights the importance of continuous surveillance to prevent infectious disease outbreaks.

Exploring Peracetic Acid and Acidic pH Tolerance of Antibiotic-Resistant Non-Typhoidal Salmonella and Enterococcus faecium from Diverse Epidemiological and Genetic Backgrounds.

Acid stress poses a common challenge for bacteria in diverse environments by the presence of inorganic (e.g., mammals' stomach) or organic acids (e.g., feed additives; acid-based disinfectants). Limited knowledge exists regarding acid-tolerant strains of specific serotypes, clonal lineages, or sources in human/animal pathogens: namely, non-typhoidal Salmonella enterica (NTS) and Enterococcus faecium (Efm). This study evaluated the acidic pH (Mueller-Hinton acidified with HCl) and peracetic acid (PAA) susceptibility of Efm (n = 72) and NTS (n = 60) from diverse epidemiological/genetic backgrounds and with multiple antibiotic resistance profiles. Efm minimum growth/survival pH was 4.5-5.0/3.0-4.0, and for NTS it was 4.0-4.5/3.5-4.0. Efm distribution among acidic pH values showed that only isolates of clade-non-A1 (non-hospital associated) or the food chain were more tolerant to acidic pH compared to clade-A1 (hospital-associated clones) or clinical isolates (p < 0.05). In the case of NTS, multidrug-resistant (MDR) isolates survived better in acidic pH (p < 0.05). The PAA MIC/MBC for Efm was 70-120/80-150 mg/L, and for NTS, it was 50-70/60-100 mg/L. The distribution of Efm among PAA concentrations showed that clade-A1 or MDR strains exhibited higher tolerance than clade-non-A1 or non-MDR ones (p < 0.05). NTS distribution also showed higher tolerance to PAA among non-MDR and clinical isolates than food chain ones (p < 0.05) but there were no differences among different serogroups. This unique study identifies specific NTS or Efm populations more tolerant to acidic pH or PAA, emphasizing the need for further research to tailor controlled measures of public health and food safety within a One Health framework.

In vitro activity of cefiderocol against Pseudomonas aeruginosa demonstrating evolved resistance to novel β-lactam/β-lactamase inhibitors.

Cefiderocol demonstrates excellent activity against MDR Pseudomonas aeruginosa; however, the activity against isolates from patients previously treated with β-lactam agents is unknown. We aimed to determine the activity of cefiderocol against P. aeruginosa collected before and after treatment with traditional β-lactams and new β-lactam/β-lactamase inhibitors. Cefiderocol MICs were determined in triplicate in iron-depleted cation-adjusted Mueller-Hinton broth and compared with β-lactam MICs tested by standard methods. All isolates underwent WGS analysis to identify mutations associated with resistance. One hundred and seventy-eight P. aeruginosa isolates were evaluated; 48% (86/178) were non-susceptible to ceftazidime/avibactam, ceftolozane/tazobactam and/or imipenem/relebactam. The cefiderocol MIC50 and MIC90 were 0.12 and 1 mg/L, respectively. Median cefiderocol MICs did not vary against isolates classified as MDR, XDR, or those non-susceptible to ceftazidime/avibactam, ceftolozane/tazobactam and/or imipenem/relebactam when compared with non-MDR isolates. Against isolates collected from patients previously treated with ceftolozane/tazobactam, cefiderocol MICs were increased 4-fold compared with baseline. Cross-resistance to cefiderocol was identified in 21% (3/14) of patients who developed treatment-emergent resistance to ceftolozane/tazobactam. Overall, 6% (11/178) of isolates demonstrated cefiderocol MICs ≥2 mg/L, which were disproportionately collected from patients previously treated with ceftolozane/tazobactam (73%; 8/11). Isolates with reduced cefiderocol susceptibility harboured mutations in ampC, tonB-dependent receptors, the response regulator pirR and ftsI. Cefiderocol demonstrates excellent in vitro activity against P. aeruginosa isolates exposed to other novel β-lactam agents; however, some exceptions were identified. Cross-resistance between cefiderocol and ceftolozane/tazobactam was evident, but not with ceftazidime/avibactam or imipenem/relebactam. Reduced cefiderocol susceptibility was mediated by mutations in ampC and tonB-dependent receptors.

A correlation study between virulence factors and multidrug resistance among clinical isolates of Proteus mirabilis

Treatment of Proteus mirabilis infections is a challenge due to the high abundance of virulence factors and the high intrinsic resistance to antimicrobials. Multidrug resistance (MDR) and extensive drug resistance (XDR) further challenge the control of P. mirabilis infection. This study aimed to investigate the correlation between virulence determinants and multidrug resistance in 100 clinical isolates of P. mirabilis collected in Alexandria from December 2019 to June 2021. Susceptibility to antimicrobials was tested by the Kirby Bauer method. Detection of swarming, urease, protease, hemolysin, and biofilm formation was performed phenotypically and by PCR amplification of zapA, flaA, ureC, mrpA, atfA, ucaA, hpmA, and luxS. MDR and XDR were detected in 34% and 5%, respectively. All isolates were positive for motility, swarming, urease, and protease production. Ninety percent were positive for hemolysin production, while 73% formed biofilm. All isolates possessed the ureC and zapA genes. The luxS, flaA, ucaA, hpmA, mrpA, and atfA genes were detected in 99%, 98%, 96% 90%, 89%, and 84%, respectively. The presence of a single biofilm-related gene was statistically correlated with non-biofilm production (P= 0.018). It was concluded that P. mirabilis isolates from catheterized-urine samples were significantly associated with biofilm formation. MDR and virulence were not statistically correlated. A significant positive correlation was detected between some virulence genes in P. mirabilis. Non-MDR isolates of P. mirabilis had a high abundance of virulence factors with no statistically significant difference from MDR. Most of the MDR and all XDR isolates could produce biofilm.

Uropathogenic Escherichia coli population structure and antimicrobial susceptibility in Norfolk, UK.

Urinary tract infections (UTIs) are a frequent cause for visits to primary care providers. In alignment globally, uropathogenic Escherichia coli (UPEC) are the main aetiological agent for UTIs in Norfolk and are increasingly difficult to treat due to multi-drug resistance. We set out to identify which clonal groups and resistance genes are disseminating in the community and hospitals in Norfolk, the first study of its kind for UPEC in this region. We collected 199 clinical E. coli isolates causing UTIs in the community and hospital from the Clinical Microbiology laboratory at Norfolk and Norwich University Hospital between August 2021 and January 2022. These were whole-genome sequenced using the Illumina and MinION platforms for in silico MLST and antibiotic resistance determinant detection. The isolates were composed of 70 STs; 8 lineages represented 56.7% of this population: ST73, ST12, ST69, ST131, ST404, ST95, ST127 and ST1193. Importantly, primary UTI screening deemed 6.5% of isolates to be multidrug resistant (MDR), with high rates of resistance to ampicillin (52.1%) and trimethoprim (36.2%) in hospitals. Of concern is the probable clonal expansion of MDR groups ST131 and ST1193 in hospitals and community settings with chromosomally encoded blaCTX-M-15, blaOXA-1 and aac(6')-Ib-cr5. The burden of reported UTIs in Norfolk is largely caused by non-MDR isolates and mirrors similar UPEC studies nationally and internationally. Continually monitoring samples with consideration of sources will help reduce burden of disease.

Pomegranate peel extract is an effective agent against MDR bacteria.

Multidrug-resistant (MDR) bacteria are one of the major public health threats facing humanity. Infections with MDR strains are difficult or impossible to treat with standard antibiotics leading to severe illnesses and even deaths. The spread of MDR bacteria has necessitated the search for alternative approaches that tackle MDR pathogens. Natural plants can be utilized as alternative therapeutic agents against the rise of MDR bacteria. In this study, we aimed to assess the antimicrobial activity of pomegranate peel extracts (PPE) against MDR clinical isolates. A total of 9 clinical isolates (8 MDR and 1 non-MDR clinical isolates) were collected and examined for their susceptibility to PPE. Using the zone of inhibition assay, 4 isolates (S. aureus, three MRSA isolates, Vancomycin-resistant Enterococci (VRE), and Acinetobacter baumannii) were sensitive to PPE. In Broth assay, 4 mg/ml PPE significantly reduced the growth (S. aureus, three MRSA isolates, Vancomycin-resistant Enterococci (VRE), and Acinetobacter baumannii), while 40 mg/ml PPE either significantly reduced or completely inhibited the growth of the isolates. The minimum bactericidal concentration (MBC) of PPE against S. aureus and MRSA-88 was 10 mg/ml. This study showed the potential of PPE as an alternative compound for treating infections caused by PPE-sensitive MDR bacteria.

Characterization of chicken eggs associated Escherichia coli and Staphylococcus aureus for biofilm production and antimicrobial resistance traits

The present study assessed the prevalence, virulence characteristics, antimicrobial resistance and biofilm-forming ability of E. coli and S. aureus recovered from egg samples in Ludhiana, Punjab. A total of 393 samples from hatcheries (n = 238), retail shops (n = 94), and households (n = 61) were collected. The prevalence of E. coli was observed as 11.70% and 9.16% for S. aureus. A total of 41.30% of E. coli isolates were positive for aggR gene and 52.17% were for fimA gene; while 36.11% of the S. aureus isolates were positive for coa gene. A high proportion of E. coli (76.10%) and S. aureus (69.44%) isolates were resistant toward ≥3 tested antibiotic classes. A total of 39.13% of E. coli isolates were moderate biofilm former, whereas the majority of the S. aureus (41.67%) were weak biofilm former. No significant difference regarding biofilm formation was observed between MDR and non-MDR isolates of E. coli and S. aureus. Biofilm genes viz., fimC and crl were reported in 43.47% and 80.43% of E. coli isolates, respectively; while icaA and icaD genes were reported in 58.34% and 47.22% of S. aureus isolates, respectively. A strong metabolic activity among 52.17% of E. coli and 41.66% of S. aureus isolates was observed using XTT assay. The present study highlights the need for applied food safety measures across the egg production chain of the region to prevent the development of MDR strains and biofilms.

Susceptibility of Salmonella serotypes isolated from meat and meat contact surfaces to thermal, acidic, and alkaline treatments and disinfectants.

The present study was conducted to evaluate the response of 29 Salmonella isolates to exposure to thermal (60°C for 2min), acidic (pH2.9 for 30 min), and alkaline (pH 11 for 60 min) treatments and investigate the susceptibility of the isolates and their biofilms to disinfectants. The reductions of Salmonella isolates populations subjected to each treatment were analyzed according to their isolation source, serotype, antibiotic resistance pattern, and biofilm formation ability. Median reductions for all of Salmonella isolates populations after thermal, acidic, and alkaline treatments were 1.8, 2.1, and 0.7 log CFU/ml, respectively. The isolates behavior under stress conditions were not related to their isolation source, serotype, or biofilm formation ability. The median reduction after alkaline treatment in non-MDR (multidrug- resistant) isolates populations was significantly (p < .05) higher than MDR isolates. The median reduction in biofilms of moderate biofilm producers by disinfectants was significantly (p < .05) higher than that of strong biofilm producers. In conclusion, Salmonella isolates showed the highest susceptibility to acidic treatment and MDR isolates were more resistant to alkaline treatment than non-MDR ones. The current study also revealed that the strong biofilm producer isolates were more resistant to disinfectants than moderate biofilm producers. This study facilitated the understanding of the relationship between Salmonella characteristics (isolation source, serotype, antibiotic resistance pattern, and biofilm formation ability) and its susceptibility to thermal, acidic, and alkaline treatments and disinfectants. The findings are helpful for the prevention and control of Salmonella.

A one-year genomic investigation of Escherichia coli epidemiology and nosocomial spread at a large US healthcare network

BackgroundExtra-intestinal pathogenic Escherichia coli (ExPEC) are a leading cause of bloodstream and urinary tract infections worldwide. Over the last two decades, increased rates of antibiotic resistance in E. coli have been reported, further complicating treatment. Worryingly, specific lineages expressing extended-spectrum β-lactamases (ESBLs) and fluoroquinolone resistance have proliferated and are now considered a serious threat. Obtaining contemporary information on the epidemiology and prevalence of these circulating lineages is critical for containing their spread globally and within the clinic.MethodsWhole-genome sequencing (WGS), phylogenetic analysis, and antibiotic susceptibility testing were performed for a complete set of 2075 E. coli clinical isolates collected from 1776 patients at a large tertiary healthcare network in the USA between October 2019 and September 2020.ResultsThe isolates represented two main phylogenetic groups, B2 and D, with six lineages accounting for 53% of strains: ST-69, ST-73, ST-95, ST-131, ST-127, and ST-1193. Twenty-seven percent of the primary isolates were multidrug resistant (MDR) and 5% carried an ESBL gene. Importantly, 74% of the ESBL-E.coli were co-resistant to fluoroquinolones and mostly belonged to pandemic ST-131 and emerging ST-1193. SNP-based detection of possible outbreaks identified 95 potential transmission clusters totaling 258 isolates (12% of the whole population) from ≥ 2 patients. While the proportion of MDR isolates was enriched in the set of putative transmission isolates compared to sporadic infections (35 vs 27%, p = 0.007), a large fraction (61%) of the predicted outbreaks (including the largest cluster grouping isolates from 12 patients) were caused by the transmission of non-MDR clones.ConclusionBy coupling in-depth genomic characterization with a complete sampling of clinical isolates for a full year, this study provides a rare and contemporary survey on the epidemiology and spread of E. coli in a large US healthcare network. While surveillance and infection control efforts often focus on ESBL and MDR lineages, our findings reveal that non-MDR isolates represent a large burden of infections, including those of predicted nosocomial origins. This increased awareness is key for implementing effective WGS-based surveillance as a routine technology for infection control.

Fosfomycin susceptibility among Escherichia coli causing urinary tract infection in a tertiary care centre in Western Maharashtra

Background and objective: Urinary tract infection(UTI) is one of the most common bacterial infections encountered in clinical practice. UTIs caused by extended-spectrum beta-lactamase (ESBL) AmpC and metallo-beta-lactamase (MBL) producing Escherichia coli (E. coli) are difficult to treat. Fosfomycin is an old antibiotic that has excellent bactericidal activity against a wide range of bacteria. This study aimed to determine the fosfomycin susceptibility of E. coli causing UTI in a tertiary care hospital in Western Maharashtra, India. Material and methods: The study was conducted at a tertiary care center in Pune, a city of Western Maharashtra, India. Urine samples from UTI cases yielding significant (&gt; 1x 105 cfu/ml) growth of E. coli were included in study. E. coli isolates were tested for susceptibility to fosfomycin and a panel of antimicrobial agents by Kirby Bauer disc diffusion method. All the isolates were tested for production of ESBL, AmpC and MBL. Result: A total of 88 E. coli were isolated of which, 47 (53.40%) and 41 (46.59%) were from male and female patients respectively. Of the total E. coli isolates, 58 (65.9%) were from in-patient cases. Multi-drug resistance was found in 69 (78.40%) isolates and remaining 19 (21.6%) were resistant to different antimicrobials tested. All (100%) the MDR and non-MDR isolates were sensitive to fosfomycin. Highest resistance was present against nalidixic acid (93.8%) while resistance was least against nitrofurantoin (15.91%), piperacillin/tazobactam (17.1%) and meropenem (18.18%). Of the total, 35 (50.72%) isolates were both AmpC and ESBL producers while 11 (15.94%) and 8 (11.59%) were only AmpC and ESBL producers respectively. MBL was positive in 15 (21.73%) of E. coli isolates. All those isolates tested sensitive to fosfomycin. Conclusion: The study revealed that fosfomycin had excellent activity against MDR E. coli causing UTI in our area. IMC J Med Sci. 2023; 17(1): 008. DOI: https://doi.org/10.55010/imcjms.17.008 *Correspondence: Dr. Sameena Khan, Department of Microbiology, Dr. D. Y. Patil Medical College, Hospital and Research Centre, Dr. D. Y. Patil Vidyapeeth, Pimpri, Pune, Maharashtra, India. E-mail: sameenak27@gmail.com

Genetic characterization of third- or fourth-generation cephalosporin-resistant avian pathogenic Escherichia coli isolated from broilers.

The third- or fourth-generation cephalosporins (3GC or 4 GC) are classified as "critically important antimicrobials for human medicine" by WHO, but resistance to these drugs is increasing rapidly in avian pathogenic E. coli (APEC). This study investigated the distribution and genetic characteristics of 3GC- or 4 GC-resistant APEC isolates from five major integrated broiler operations in Korea. The prevalence of 3GC- or 4GC-resistant APEC isolates in 1-week-old broilers was the highest in farms of operation C (53.3%); however, the highest prevalence of these isolates in 4-week-old broilers was the highest on the farms of operation A (60.0%), followed by operations E (50.0%) and C (35.7%). All 49 3GC- or 4GC-resistant APEC isolates had at least one β-lactamase-encoding gene. The most common β-lactamase-encoding genes was extended-spectrum β-lactamase gene, bla CTX-M-15, detected in 24 isolates (49.0%), followed by bla TEM-1 (32.7%). Sixteen isolates (32.7%) harbored class 1 integrons, and four isolates (8.2%) showed different gene cassette-arrangements. However, only 1 of 26 isolates harboring class 2 integrons carried a gene cassette. Furthermore, both CRISPR 1 and 2 arrays were detected in most isolates (36 isolates; 73.5%), followed by CRISPR 2 (18.4%) and CRISPR 1 (4.1%). Interestingly, CRISPR 2 was significantly more prevalent in multidrug resistant (MDR)-APEC isolates than in non-MDR APEC isolates, whereas CRISPR 3 and 4 were significantly more prevalent in non-MDR APEC isolates (each 11.1%; p < 0.05). None of the protospacers of CRISPR arrays were directly associated with antimicrobial resistance. Our findings indicate that the distribution and characteristics of 3GC or 4GC-resistant APEC isolates differed among the integrated broiler operations; moreover, improved management protocols are needed to control the horizontal transmission of 3GC or 4GC-resistant APEC isolates.

Acinetobacter baumannii Biofilm Formation: Association with Antimicrobial Resistance and Prolonged Survival under Desiccation

Biofilm-forming multidrug-resistant Acinetobacter baumannii has emerged as a global pathogen. This study investigated the impact of biofilm formation by A. baumannii on antimicrobial resistance and prolonged survival under desiccation, which is essential for effective infection control of A. baumannii in hospital settings. Seventy-eight clinical isolates of A. baumannii were identified, and antibiotic susceptibility profiles were assessed. All the isolates were investigated for their biofilm-forming abilities at 24 and 48h. The biofilm inhibitory concentrations of antibiotics were evaluated for selected biofilm-forming isolates to determine the influence of biofilm on antibiotic tolerance. The impact of biofilm formation on desiccation tolerance was also evaluated for up to 48days. The results revealed that out of 78 A. baumannii clinical isolates, 83% were MDR and 17% non-MDR. Overall, 79% of isolates formed high biofilm after 24h. The extent of biofilm formation gets significantly increased after 48h, and 87% of isolates formed high biofilm. It was observed that eradicating mature biofilm requires up to a thousandfold higher concentration of antibiotics than MICs, and biofilm-forming isolates can survive for a prolonged period under desiccation. In conclusion, our findings revealed that both MDR and non-MDR isolates of A. baumannii could form biofilms on abiotic surfaces. A. baumannii biofilms contribute to endurance in the presence of antimicrobials and desiccation conditions, which are significant trouble for hospital patient care management. The present findings may offer insights for developing preventive measures to tackle biofilm-associated A. baumannii infection.

Multidrug-resistant Pseudomonas aeruginosa is predisposed to lasR mutation through up-regulated activity of efflux pumps in non-cystic fibrosis bronchiectasis patients.

BackgroundMultidrug-resistant (MDR) Pseudomonas aeruginosa is a frequent opportunistic pathogen that causes significant mortality in patients with non-cystic fibrosis bronchiectasis (NCFB). Although the quorum sensing (QS) system is a potential target for treatment, lasR mutants that present with a QS-deficient phenotype have been frequently reported among clinical P. aeruginosa isolates. We aimed to investigate whether antibiotic resistance would select for lasR mutants during chronic P. aeruginosa lung infection and determine the mechanism underlying the phenomenon.MethodsWe prospectively evaluated episodes of chronic P. aeruginosa lung infections in NCFB patients over a 2-year period at two centers of our institution. QS phenotypic assessments and whole-genome sequencing (WGS) of P. aeruginosa isolates were performed. Evolution experiments were conducted to confirm the emergence of lasR mutants in clinical MDR P. aeruginosa cultures.ResultsWe analyzed episodes of P. aeruginosa infection among 97 NCFB patients and found only prior carbapenem exposure independently predictive of the isolation of MDR P. aeruginosa strains. Compared with non-MDR isolates, MDR isolates presented significantly QS-deficient phenotypes, which could not be complemented by the exogenous addition of 3OC12-HSL. The paired isolates showed that their QS-phenotype deficiency occurred after MDR was developed. Whole-genome sequencing analysis revealed that lasR nonsynonymous mutations were significantly more frequent in MDR isolates, and positive correlations of mutation frequencies were observed between genes of lasR and negative-efflux-pump regulators (nalC and mexZ). The addition of the efflux pump inhibitor PAβN could not only promote QS phenotypes of these MDR isolates but also delay the early emergence of lasR mutants in evolution experiments.ConclusionsOur data indicated that MDR P. aeruginosa was predisposed to lasR mutation through the upregulated activity of efflux pumps. These findings suggest that anti-QS therapy combined with efflux pump inhibitors might be a potential strategy for NCFB patients in the challenge of MDR P. aeruginosa infections.