In vitro antimicrobial susceptibility of clinical respiratory isolates to ceftazidime-avibactam and comparators (2016-2018).

BACKGROUND: This antimicrobial surveillance study reports in vitro antimicrobial activity and susceptibility data for a panel of agents against respiratory isolates of Enterobacterales and Pseudomonas aeruginosa. METHODS: Isolates from respiratory specimens were collected in Africa/Middle East, Asia/South Pacific, Europe and Latin America between 2016 and 2018, as part of the Antimicrobial Testing Leadership and Surveillance (ATLAS) program. Broth microdilution methodology was used to quantify minimum inhibitory concentrations, from which rates of susceptibility were determined using EUCAST breakpoints (version 10). Rates of subsets with genes encoding ß-lactamases (extended-spectrum ß-lactamases [ESBLs], serine carbapenemases and metallo-ß-lactamases [MBLs]) were also determined, as well as rates of multidrug-resistant (MDR) P. aeruginosa. RESULTS: Among all respiratory Enterobacterales isolates, susceptibility to ceftazidime-avibactam, meropenem, colistin and amikacin was ≥94.4% in each region. For Enterobacterales isolates that were ESBL-positive or carbapenemase-positive/MBL-negative, ceftazidime-avibactam susceptibility was 93.6 and 98.9%, respectively. Fewer than 42.7% of MBL-positive Enterobacterales isolates were susceptible to any agents, except colistin (89.0% susceptible). Tigecycline susceptibility was ≥90.0% among Citrobacter koseri and Escherichia coli isolates, including all ß-lactamase-positive subsets. ESBL-positive Enterobacterales were more commonly identified in each region than isolates that were ESBL/carbapenemase-positive; carbapenemase-positive/MBL-negative; or MBL-positive. Among all respiratory P. aeruginosa isolates, the combined susceptibility rates (susceptible at standard dosing regimen plus susceptible at increased exposure) were highest to ceftazidime-avibactam, colistin and amikacin (≥82.4% in each region). Susceptibility to colistin was ≥98.1% for all ß-lactamase-positive subsets of P. aeruginosa. The lowest rates of antimicrobial susceptibility were observed among MBL-positive isolates of P. aeruginosa (≤56.6%), with the exception of colistin (100% susceptible). MDR P. aeruginosa were most frequently identified in each region (18.7-28.7%), compared with the subsets of ESBL-positive; carbapenemase-positive/MBL-negative; or MBL-positive isolates. CONCLUSIONS: Rates of susceptibility among the collections of respiratory Enterobacterales and P. aeruginosa isolates were highest to ceftazidime-avibactam, colistin and amikacin in each region. Tigecycline was active against all subsets of C. koseri and E. coli, and colistin was active against all subsets of P. aeruginosa. The findings of this study indicate the need for continued antimicrobial surveillance among respiratory Gram-negative pathogens, in particular those with genes encoding MBLs.

Genome sequence of carbapenem-resistant Citrobacter koseri carrying blaOXA-181 isolated from sewage sludge.

OBJECTIVES: This study reported the resistome content of sewage sludge-isolated carbapenem-resistant Citrobacter koseri (C. koseri) carrying blaOXA-181. It also provided a general phylogenomic analysis highlighting antibiotic resistance genes (ARGs), plasmids and pathogenicity of C. koseri genomes. METHODS: The carbapenem-resistantC. koseri AS1 strain was isolated from sewage sludge on CHROMagar™ mSuperCARBA™ media. Whole genome sequencing of C. koseri AS1 was performed using an HiSeq X™ Ten instrument. Additional C. koseri genomes were downloaded from National Center for Biotechnology Information (NCBI). Phylogenomic analysis was established through CSI Phylogeny. ARGs, plasmids and pathogenicity were identified using ResFinder 3.1, PlasmidFinder 2.0 and PathogenFinder 1.1, respectively. RESULTS: The phylogenomic tree indicated a polyclonal pattern ofC. koseri genomes. Resistome analysis of C. koseri AS1 revealed ß-lactam resistance genes (blaMAL-1 and blaOXA-181) as well as a fosfomycin resistance gene (fosA7). Three plasmids (ColKP3, ColRNAI and IncX30) were identified in the C. koseri AS1 genome. In addition, 25 ARGs were found in downloaded genomes. Of these, clinically significant ARGs such as blaKPC-2 and blaOXA-48 were found in two and four genomes, respectively. Assessment of the genomes using PathogenFinder revealed all genomes as putative human pathogens. CONCLUSIONS: It is believed that noC. koseri genome has been reported to carry blaOXA-181; therefore, C. koseri AS1 is the first of its kind. This study also highlighted the resistome contents of C. koseri genomes.

Genotypic study of Citrobacter koseri, an emergent platelet contaminant since 2012 in France.

BACKGROUND: Transfusion-transmitted bacterial infection is a rare occurrence but the most feared complication in transfusion practices. Between 2012 and 2017, five cases of platelet concentrates (PCs) contaminated with the bacterial pathogen Citrobacter koseri (PC-Ck) have been reported in France, with two leading to the death of the recipients. We tested the possibilities of the emergence of a PC-specific clone of C. koseri (Ck) and of specific bacterial genes associated with PC contamination. STUDY DESIGN AND METHODS: The phylogenetic network, based on a homemade Ck core genome scheme, inferred from the genomes of 20 worldwide Ck isolates unrelated to PC contamination taken as controls (U-Ck) and the genomes of the five PC-Ck, explored the clonal relationship between the genomes and evaluated the distribution of PC-Ck throughout the species. Along with this core genome multilocus sequence typing approach, a Ck pan genome has been used to seek genes specific to PC-Ck isolates. RESULTS: Our genomic approach suggested that the population of C. koseri is nonclonal, although it also identified a cluster containing three PC-Ck and eight U-Ck. Indeed, the PC-Ck did not share any specific genes. CONCLUSION: The elevated incidence of PCs contaminated by C. koseri in France between 2012 and 2017 was not due to the dissemination of a clone. The determinants of the recent outbreaks of PC contamination with C. koseri are still unknown.

The Effect of Ten Essential Oils on Several Cutaneous Drug-Resistant Microorganisms and Their Cyto/Genotoxic and Antioxidant Properties.

In this study, we determined the antimicrobial activity of ten essential oils (EOs)-oregano, thyme, clove, arborvitae, cassia, lemongrass, melaleuca, eucalyptus, lavender, and clary sage-against drug-resistant microorganisms previously isolated from patients with skin infections. The essential oil compositions were determined using gas chromatography coupled to mass spectrometry (GC/MS). The assayed bacteria included Pseudomonas aeruginosa, Proteus vulgaris, Citrobacter koseri, and Klebsiella pneumoniae. Two drug-resistant yeasts (Candida albicans and Candida parapsilosis) were also involved in our survey. Oregano, thyme, cassia, lemongrass and arborvitae showed very strong antibacterial and antifungal activity against all tested strains. These results show that these essential oils may be effective in preventing the growth of the drug-resistant microorganisms responsible for wound infections. In this study, the genotoxic effects of tested essential oils on healthy human keratinocytes HaCaT were evaluated using the comet assay for the first time. These results revealed that none of the essential oils induced significant DNA damage in vitro after 24 h. Moreover, the treatment of HaCaT cells with essential oils increased the total antioxidant status (TAS) level. The obtained results indicate that EOs could be used as a potential source of safe and potent natural antimicrobial and antioxidant agents in the pharmaceutical and food industries.

First genome sequence of a blaKPC-2-carrying Citrobacter koseri isolate collected from a patient with diarrhoea.

OBJECTIVES: The blaKPC gene is rarely reported in Citrobacter koseri. Here we report the first draft genome sequence of a blaKPC-2-carrying C. koseri isolate from a patient with diarrhoea. METHODS: Transferability of the blaKPC-2-bearing plasmid was determined by the filter mating method. The whole genome sequence of C. koseri L168 was determined using an Illumina HiSeq platform. The genome was de novo assembled using Velvet 1.2.10. Acquired antimicrobial resistance genes and plasmid replicons were identified using ResFinder 2.1 and PlasmidFinder 1.3, respectively. RESULTS: Antimicrobial susceptibility testing (AST) showed that C. koseri L168 was resistant to multiple antibiotics but was susceptible to ciprofloxacin, gentamicin, tobramycin, amikacin, tigecycline and colistin. A KPC-2-harbouring plasmid was conjugative and the transconjugants conferred increased resistance to carbapenems confirmed by conjugation experiments and AST. In silico analysis revealed the presence of the ß-lactam resistance genes blaKPC-2 and blaMAL-1. Additionally, plasmids of incompatibility groups IncFII and IncX4 were identified in the genome by PlasmidFinder. BLAST analysis revealed that blaKPC-2 was located on a Tn3 transposon element in C. koseri L168 with the conserved linear structure ISKpn27-blaKPC-2-ΔISKpn6-korC-klcA. CONCLUSIONS: To our knowledge, this is only the second report of C. koseri producing KPC-2, and we report the first draft genome sequence of a blaKPC-2-carrying C. koseri isolate from a patient with diarrhoea in China. This work may facilitate our understanding of the pathogenesis, multidrug resistance mechanisms and genomic features of this species. Further monitoring of bacteria carrying carbapenemase genes in patients' gut microbiota is warranted.

Efficacy and safety of 3 day versus 7 day cefditoren pivoxil regimens for acute uncomplicated cystitis: multicentre, randomized, open-label trial.

BACKGROUND: Fluoroquinolone-non-susceptible Escherichia coli isolated from patients with acute uncomplicated cystitis are a matter of increasing concern. Cefditoren pivoxil is an oral, ß-lactamase-stable, extended-spectrum cephalosporin that is effective against fluoroquinolone-non-susceptible bacteria. OBJECTIVES: To evaluate the clinical and microbiological efficacies of cefditoren pivoxil against acute uncomplicated cystitis and to determine the optimal duration of cefditoren pivoxil treatment. METHODS: We compared 3 and 7 day regimens of cefditoren pivoxil in a multicentre, randomized, open-label study. RESULTS: A total of 104 female patients with acute uncomplicated cystitis were enrolled and randomized into 3 day (n = 51) or 7 day (n = 53) treatment groups. At first visit, 94 bacterial strains were isolated from the 104 participants of which 81.7% (85/104) were E. coli. Clinical and microbiological efficacies were evaluated 5-9 days following administration of the final dose of cefditoren pivoxil. The clinical efficacies of the 3 and 7 day groups were 90.9% (40/44) and 93.2% (41/44), respectively (P = 1.000). The microbiological efficacies of the 3 and 7 day groups were 82.5% (33/40) and 90.2% (37/41), respectively (P = 0.349). There were no adverse events due to cefditoren pivoxil treatment, with the exception of a mild allergic reaction in one patient, after which the cefditoren pivoxil was exchanged for another antimicrobial. CONCLUSIONS: Cefditoren pivoxil is safe and effective for uncomplicated cystitis, with no significant differences in clinical and microbiological efficacies between 3 and 7 day regimens.

blaNDM-1 Carriage on IncR Plasmid in Enterobacteriaceae Strains.

Four NDM-1-producing Enterobacteriaceae strains (three Klebsiella pneumoniae and one Citrobacter koseri) were isolated between 2009 and 2011 through a nationwide surveillance for carbapenem-resistant Enterobacteriaceae in Croatia to study the molecular genetic background of blaNDM and the responsible plasmid types. Phenotypically, the clinical strains proved to be multidrug resistant. All strains remained susceptible to tigecycline and colistin. The clinical strains harbored variable antibiotic resistance determinants, notably, blaNDM-1, blaTEM-1, blaSHV-1, blaSHV-12, blaOXA-1, blaOXA-9, blaCTX-M-15, blaCMY-4, qnrB1, and aac(6')Ib-cr in different combinations. Two K. pneumoniae belonged to sequence type ST15 and one strain to ST16. As for the plasmid types, C. koseri and one of the ST15 K. pneumoniae carried IncR, and the second ST15 K. pneumoniae carried IncR and colE. The K. pneumoniae ST16 strain hosted A/C and colE plasmids. The blaNDM-1 gene was detected on conjugative high-molecular-weight plasmids, namely, A/C and IncR types. It is noteworthy that this is the first description of K. pneumoniae ST16 expressing NDM-1 in Europe. Remarkably, our study underscores the importance of the IncR plasmid as a reservoir of multidrug resistance. To the best of our knowledge, the IncR plasmid carrying blaNDM-1 in C. koseri is reported for the first time.

Detection of an IncA/C plasmid encoding VIM-4 and CMY-4 ß-lactamases in Klebsiella oxytoca and Citrobacter koseri from an inpatient in a cardiac rehabilitation unit.

A 62-year-old patient was transferred to the cardiac rehabilitation unit of the I.R.C.C.S. Fondazione S. Maugeri after undergoing a heart transplantation at the Acute Care Hospital I.R.C.C.S. S. Matteo of Pavia. On 1 August 2013 and during hospitalization in the rehabilitation unit, Klebsiella oxytoca and Citrobacter koseri clinical isolates were simultaneously recovered from the patient's preputial swab. Both the K. oxytoca and C. koseri strains were carbapenem- resistant by MicroScan System (Beckman Coulter). Carbapenem-resistant K. pneumoniae had previously been reported in the same rehabilitation facility. The aim of the study was to identify the carbapenem resistance mechanisms among the enterobacterial species recovered. Phenotypic screening tests useful to detect the ß-lactamases/carbapenemases were performed. Carbapenem MICs were obtained by Etest. AmpC and MBL encoding genes were identified by PCR and sequencing. Conjugation assays and plasmid characterization were performed. Both of the K. oxytoca and C. koseri isolates were multi drug resistant, showing resistance to amoxicillin-clavulanic acid, three generation cephalosporins, ertapenem (K. oxytoca MIC, >32 mg/L; C. koseri MIC, 4 mg/L), imipenem (K. oxytoca MIC, 4 mg/L; C. koseri MIC, 12 mg/L), thrimethoprim sulphamethoxazole and gentamicin. Susceptibility was retained to fluoroquinolones, colistin and tigecycline. Molecular characterization confirmed the co-presence of blaCMY-4 and blaVIM-4 determinants in a 150 Kb transferable plasmid of IncA/C group. This case is the first detection in Italy of the K. oxytoca and C. koseri clinical isolates co-producing the CMY-4 and VIM-4 enzymes.

Optimum management of Citrobacter koseri infection.

Low virulent Citrobacter koseri can cause life threatening infections. Neonates and other immunocompromised patients are particularly susceptible to infection from C. koseri. Any infection due to C. koseri mandates antimicrobial therapy based on the sensitivity of the pathogen microorganism. Various types of antibiotics, including aminoglycosides carbapenems, cephalosporins, chloramphenicol and quinolones, are used for the treatment of C. koseri infections. The rational choice of antimicrobial therapy for Citrobacter infections is a challenge for clinicians because there is a sustained increase in antibacterial resistance. We reviewed antimicrobial agents used for C. koseri infections in this review.

Iron induces bimodal population development by Escherichia coli.

Bacterial biofilm formation is a complex developmental process involving cellular differentiation and the formation of intricate 3D structures. Here we demonstrate that exposure to ferric chloride triggers rugose biofilm formation by the uropathogenic Escherichia coli strain UTI89 and by enteric bacteria Citrobacter koseri and Salmonella enterica serovar typhimurium. Two unique and separable cellular populations emerge in iron-triggered, rugose biofilms. Bacteria at the air-biofilm interface express high levels of the biofilm regulator csgD, the cellulose activator adrA, and the curli subunit operon csgBAC. Bacteria in the interior of rugose biofilms express low levels of csgD and undetectable levels of matrix components curli and cellulose. Iron activation of rugose biofilms is linked to oxidative stress. Superoxide generation, either through addition of phenazine methosulfate or by deletion of sodA and sodB, stimulates rugose biofilm formation in the absence of high iron. Additionally, overexpression of Mn-superoxide dismutase, which can mitigate iron-derived reactive oxygen stress, decreases biofilm formation in a WT strain upon iron exposure. Not only does reactive oxygen stress promote rugose biofilm formation, but bacteria in the rugose biofilms display increased resistance to H(2)O(2) toxicity. Altogether, we demonstrate that iron and superoxide stress trigger rugose biofilm formation in UTI89. Rugose biofilm development involves the elaboration of two distinct bacterial populations and increased resistance to oxidative stress.

Detection of Citrobacter koseri carrying beta-lactamase KPC-2 in a hospitalised patient, Greece, July 2011.

This report describes the detection of Citrobacter koseri carrying K. pneumoniae carbapenemase (KPC-2) isolated in July 2011 from a Greek patient, who was also colonised by a Klebsiella pneumoniae strain coproducing KPC-2 and Verona integron-encoded metallo-beta-lactamase (VIM)-1.

Comparison of beta-lactamase genes in clinical and food bacterial isolates in India.

BACKGROUND: The present study aimed to determine the occurrence of human disease-causing enteric bacteria on raw vegetables, fruits, meats, and milk products sold in Indian markets. The study further aimed to analyze antibiotic resistance rates and the presence of blaCTX-M, blaTEM, blaSHV, and blaAmpC. METHODOLOGY: Twenty-three food-borne and 23 clinical isolates were compared for antibiotic resistance rates and the presence of blaCTX-M, blaTEM, blaSHV, and blaAmpC. Swabs were taken from unwashed and washed food items, as well as from some chopped food specimens, and inoculated on appropriate culture medium. Bacterial isolates were identified, antibiotic susceptibility was performed, and bla genes were detected by PCR. RESULTS: Thirty-eight bacterial isolates were obtained from the food specimens, of which 36 (94.7%) were Gram-negative and two (5.3%) were Gram-positive bacterial species. Klebsiella pneumoniae was the most prevalent (52.6%; 20/38) bacterial species isolated, followed by Citrobacter koseri (18.4%; 7/38). In food isolates, the majority of the isolates were resistant to gentamicin (33.3%) followed by amikacin (11.1%). Resistance to a third-generation cephalosporin was noticed in only 5.6% isolates. However, in clinical isolates, maximal resistance was noticed against third-generation cephalosporins followed by ofloxacin in 91.3% and 86.9% isolates, respectively, and resistance to gentamicin and amikacin was noticed in 78.3% and 52.2% isolates, respectively. The presence of blaCTX-M, blaTEM, blaSHV, and blaAmpC in clinical isolates was noticed in 52.2%, 60.9%, 21.7%, and 43.5%, respectively. None of the isolates from food showed the presence of any of the above-cited genes. CONCLUSIONS: Probably bla genes have not yet disseminated to raw-food vegetation in India.

Treatment of Citrobacter koseri infection with ciprofloxacin and cefotaxime in a preterm infant.

OBJECTIVE: To report a case of successful treatment of Citrobacter koseri infection in a preterm infant as a means of challenging the current treatment recommendations on the basis of pharmacodynamic and pharmacokinetic considerations. CASE SUMMARY: A premature infant was diagnosed with C. koseri sepsis after 3 weeks in intensive care. Concern for meningitis was based on the propensity for central nervous system (CNS) involvement with Citrobacter infection along with new findings of ventriculomegaly and hydrocephalus shown on cranial ultrasound (CUS). The infant was treated with ciprofloxacin 10-20 mg/day and cefotaxime 100 mg/day for 21 days. After treatment, lumbar puncture was normal, follow-up CUS returned to baseline, and the infant passed a hearing screen after discharge. A favorable outcome was achieved in this case. DISCUSSION: Approximately 76% of neonatal patients infected with C. koseri develop brain abscesses. The mortality rate for meningitis due to Citrobacter spp. is approximately 30%, and of the infants who survive, more than 80% have some degree of mental retardation. Third-generation cephalosporins and aminoglycosides are traditional therapies against this infection. The current antibiotic strategies have failed to prevent the high rates of morbidity and mortality associated with Citrobacter infections. A possible basis for these poor outcomes is failure to apply appropriate pharmacokinetic and pharmacodynamic principles in selecting antibiotics that will achieve adequate concentrations to kill the bacteria in granulocytes within the CNS. Based on favorable sensitivity data, penetration into neutrophils and the CNS, and favorable toxicity profiles, ciprofloxacin and meropenem would appear to be the most appropriate antibiotic treatment options for systemic infection or meningitis caused by C. koseri. CONCLUSIONS: Ciprofloxacin and meropenem should be considered antibiotic treatment options for systemic infection or meningitis caused by C. koseri.

A rare case of lethal retroperitoneal abscess caused by Citrobacter koseri.

Retroperitoneal abscesses are very uncommon clinical conditions. The characteristically vague symptomatology of retroperitoneal abscess and the inherent difficulty of identifying retroperitoneal disease by physical examination contributed to these dismal therapeutic outcomes. We present an unusual case of lethal retroperitoneal abscess, caused by Citrobacter diversus(koseri), treated with surgical drainage. Citrobacter species have rarely been involved in deep tissue infection and there is no reported case of lethal retroperitoneal abscess caused by C. koseri. This case is the only reported case of C. koseri as the sole pathogen associated with a lethal retroperitoneal abscess in immunocompetent patient. The case is also notable because it confirms the recent bacterial resistance to beta-lactam antibiotics and to other antimicrobial agents, like chloramphenicol or cotrimoxazol.

Citrobacter infections in a tertiary care hospital in Northern India.

OBJECTIVES: This prospective study was carried out to look for the frequency of isolation of Citrobacter species from clinical specimens and study their antimicrobial susceptibility pattern. METHODS: Patients from whom Citrobacter species were isolated during routine diagnostic testing from January to December 2004 were included in the study. Isolates were identified by standard biochemical tests. Antimicrobial susceptibility testing was performed by disk diffusion method as per National Committee of Clinical Laboratory Standards (NCCLS) guidelines. RESULTS: Citrobacter species were isolated from a total of 205 patients. Infection was nosocomially acquired in 94.6% patients. One hundred eighty one (88.3%) patients had significant underlying illnesses. Culture yielded Citrobacter koseri in 185 (90.2%) and Citrobacter freundii in 20 (9.8%) patients. The distribution of isolates was as follows: urine (46.2%), respiratory tract (16.3%), blood (15.8%), pus (12.1%) and sterile body fluids (9.3%). Drug resistance was observed to be more in C. koseri as compared to C. freundii. CONCLUSIONS: Citrobacter infections are often nosocomially acquired, seen in patients with significant underlying diseases and isolates are commonly drug-resistant. Adoption of hospital infection control practices and a good antibiotic policy may prevent their spread.

Characterization of the chromosomal class A beta-lactamase CKO from Citrobacter koseri.

The gene bla(CKO) encoding the chromosomal class A beta-lactamase of Citrobacter koseri was cloned and sequenced. CKO was found to display only 41% identity with SED-1 from Citrobacter sedlakii and 36% with CdiA from Citrobacter amalonaticus (formerly Citrobacter diversus). No transcriptional regulator was found upstream from bla(CKO). Silent and missense mutations were detected in four bla(CKO) genes amplified from different C. koseri clinical isolates, but the CKO variants displayed identical biochemical behaviours. A bla(CKO)-specific polymerase chain reaction confirmed that bla(CKO) is present only in C. koseri and therefore represents an interesting tool with which to differentiate C. koseri from the other Citrobacter spp.

Novel TEM-type extended-spectrum beta-lactamase, TEM-134, in a Citrobacter koseri clinical isolate.

A new natural TEM derivative with extended-spectrum beta-lactamase activity, TEM-134, was identified in a ceftazidime-resistant clinical isolate of Citrobacter koseri. Compared to TEM-1, TEM-134 contains the following mutations: Q39K, E104K, R164H, and G238S. The bla(TEM-134) gene was not transferable by conjugation and, apparently, was chromosomally encoded. Expression studies with Escherichia coli revealed efficient cefotaximase and ceftazidimase activity for TEM-134.