Differences In Antibiotic Susceptibility Research Articles

Development of a multiyear pediatric antibiogram in Georgia identifies antibiotic resistance trends

Background: Antibiograms are used to monitor antibiotic resistance trends and help guide empiric antibiotic treatment. Community pediatricians may not have access to or be comfortable using children’s hospital antibiograms. Creating and disseminating a statewide pediatric antibiogram can help inform antibiotic stewardship efforts.Objective: To develop a pediatric-specific antibiogram for the state of Georgia. Methods: Annual pediatric antibiograms for the 5 children’s hospitals in Georgia from 2014 through 2021 were collected. All sites complied with the Clinical and Laboratory Standards Institute guidelines for antibiomicrobic breakpoints and antibiogram development. Antibiogram data were combined, and the most common bacteria were selected to incorporate into the statewide antibiogram: Staphylococcus aureus, Streptococcus pneumoniae, Enterococcus faecalis, Escherichia coli, Klebsiella pneumoniae, Enterobacter cloacae complex, and Pseudomonas aeruginosa. Antibiogram data were reported as percentage susceptible and total number of isolates. Interhospital susceptibility differences were compared for methicillin-susceptible S. aureus (MSSA), methicillin-resistant S. aureus (MRSA), E. coli, and K. pneumoniae from 2018 through 2021. P < .05 was considered significant. The combined antibiogram data from 2014 through 2021 were used to show antibiotic susceptibility trends over time. Results: The 2021 antibiogram is shown in the Table. For MSSA and MRSA, clindamycin susceptibility was 80% and 85%, respectively. K. pneumoniae susceptibility to amoxicillin-clavulanate was 91%. For E. coli, using urine-specific breakpoints, susceptibility to cefazolin was 89%. A few statistically significant differences in antibiotic susceptibility were detected between hospitals, but most were unlikely to be clinically relevant (all susceptibilities ≥90% or < 80%). A notable exception was trimethoprim-sulfamethoxazole susceptibility for K. pneumoniae, which ranged from 74% to 98% in 2020 and from 74% to 86% in 2021. From 2014 to 2021, the percentage of MRSA decreased from 49% to 34%. Over the 8 years, susceptibility to ceftriaxone for E. coli ranged from 93% to 95% and from 90% to 95% for K. pneumoniae. Susceptibility to meropenem for E. coli and K. pneumoniae ranged from 99% to 100%. Conclusions: Antibiotic susceptibility for pediatric bacterial isolates in Georgia remained stable over time and supported the narrow-spectrum empiric antibiotic treatment recommended in national evidence-based guidelines for skin and soft-tissue infections, community-acquired pneumonia, and uncomplicated urinary tract infections. MRSA rates decreased over time and multidrug-resistant gram-negative bacilli were uncommon and remained stable.Disclosures: None

Investigation of Virulence Genes and Carbapenem Resistance Genes in Hypervirulent and Classical Isolates of Klebsiella pneumoniae Isolated from Various Clinical Specimens

Klebsiella pneumoniae is a global pathogen that can cause hospital-acquired and community-acquired infections and is known for its resistance to antibiotics. The pathotype, which is defined as hypervirulent K.pneumoniae (hvKp) is more lethal than classical K.pneumoniae (cKp) isolates and causes many community-acquired infections such as liver abscess, endophthalmitis, pneumonia in healthy young adults. There are no clear clinical or microbiological criteria to define hvKp. String test showing hypermucoviscosity and the iucA gene encoding aerobactin, a siderophore, were used to demonstrate hypervirulence. In this study, it was aimed to investigate the presence of various virulence genes and carbapenem resistance genes in the isolates of K.pneumoniae isolated from various clinical samples in our laboratory and classified as classical and hypervirulent by string test and also to detect the presence of various virulence and carbapenem resistance genes in hvKp isolates. Presence of four virulence genes (fimH-1, rmpA, magA, iucA), K1-K2 serotypes in all isolates and five carbapenem resistance genes (blaOXA-48, blaKPC, blaIMP, blaVIM, blaNDM-1) in carbapenem resistant isolates were investigated with polymerase chain reaction (PCR) method. Forty-five percent of the isolates were defined as hvKp and 55% as cKp. The fimH-1 gene was found to be positive in 94% of the isolates, the iucA gene in 37%, the magA gene (K1) in 34%, the rmpA gene in 5%, and the K2 serotype in 3% of the isolates. iucA gene was positive in 68.9% of hvKp isolates and 10.9% of cKp isolates, and the presence of iucA gene in hvKp isolates was statistically significant compared to cKp isolates (p<0.05). magA gene and K1 serotype were detected in 28.9% of hvKp isolates and 38.2% of cKp isolates. Although the magA gene ratio was high in cKp isolates, this difference was not statistically significant (p> 0.05). fimH-1 gene was found positive in 93.3% of hvKp isolates and 94.5% of cKp isolates. The rmpA gene was positive in 8.9% of hvKp isolates and 1.8% of cKp isolates. The K2 serotype was positive in 4.4% of hvKp isolates and 1.8% of cKp isolates. Although there was no statistical difference in antibiotic susceptibility between hvKp and cKp isolates; ceftriaxone, cefuroxime, ceftazidime, amikacin, cefoxitin, ertapenem, cefuroxime axetil were found to be more sensitive in hvKp isolates. Ciprofloxacin and trimethoprim sulfamethoxazole were found to be more sensitive in hvKp isolates than cKp isolates, and the difference was statistically significant (p<0.05). Although gentamicin, amoxicillin, piperacillintazobactam were not statistically significant in the cKp group, they were more sensitive than the hvKp group (p> 0.05). Carbapenem resistance were found to be 65.7% in cKp and 34.3% in hvKp isolates. Although not statistically significant, hvKp isolates were found to be more sensitive to carbapenems. The most common gene among 35 carbapenem resistant isolates was blaOXA-48 detected in 29 isolates. While the blaKPC gene was detected in five isolates, blaIMP, blaVIM and blaNDM-1 were not detected in any isolates. Sixty nine percent of blaOXA-48 positive samples were found to be in cKp isolates and 31% in hvKp isolates. It was determined that all of the blaKPC positive isolates were hvKp isolates. It was concluded that the string test and virulence factors alone would not be sufficient to show hypervirulence, and that more than one virulence factor combination should be shown in the presence of clinical features of hypervirulent infections to show hypervirulence.

How does gender affect ureaplasma and mycoplasma growth and antimicrobial susceptibility rates?

The aim of this study was to assess the presence of M. hominis and U. urealyticum agents, their distribution between male and female, and differences in antibiotic susceptibility in samples sent from Hisar Intercontinental Hospital's various clinics with the preliminary diagnosis of genitourinary system infection. The Mycoplasma IES test was used to identify M. hominis and U. urealyticum, and to determine antibiotic susceptibility results, in samples taken from patients. The findings of mycoplasma and ureaplasma culture testing samples requested between 2014 and 2021 were evaluated retrospectively from our records. M. hominis was found to be positive in 7.37% of the examinations, U. urealyticum was found to be positive in 34.98% of the examinations, and either of them were found to be positive in 22.01% of the examinations. The growth rate of M. hominis and/or U. urealyticum was determined to be 24.95% in females and 10.13% in males, with the growth rate in females being greater and statistically significant (p < 0.001). According to the antibiotic susceptibility test results, clarithromycin (R 17.91%) was the most susceptible antibiotic overall for both microorganisms, while clindamycin (R 90.28%) was the most resistant. Depending on the sex, clarithromycin (R 18.40%) was found to be the most susceptible antibiotic in females, and levofloxacin (R 10.87%) to be the most susceptible in males. Given the presence of M. hominis and U. urealyticum infections, especially in the presence of risky conditions such as pregnancy, laboratory tests for the diagnosis of these agents should be used in such studies since no urogenital infections were detected in the routine cultures of the patients followed up with the suspicion of urogenital infection. Gender differences should also be considered as a parameter in the preference of antibiotics.

A COMPARATIVE STUDY OF ANTIMICROBIAL SUSCEPTIBILITY OF E. COLI AMONG NONELDERLY VERSUS ELDERLY PATIENTS WITH URINARY TRACT INFECTION IN A TERTIARY CARE HOSPITAL.

Introduction: As there is rise in the elderly population, there is increase in the geriatric illnesses, urinary tract infection is one of the commonly seen among them. There is increasing prevalence of antibiotic resistance in the elderly patients. Aim: Study was conducted to look for the difference in antibiotic susceptibility in nonelderly compared to elderly patients. We took 100 cases with 50 elderly and 50 nonelderly with E.coli grown in urine Material and methods: samples with features of urinary tract infection. There was no significant difference in the antibi Results: otic susceptibility in both the groups except for the cefepime being good choice in nonelderly amongst cephalosporins. Amongst oral antibiotics to choose empirically are Nitrofurantoin, Cotrimoxazole and Amoxycillin in both the groups with no significant differences. We can use all the groups of antibiotics empirically especially inj Conclusions: ectables like Cephalosporins, Piperacillin, Aminoglycosides and Carbapenems as and when indicated in both groups with almost equal results. But we need to take a prospective study with larger number of cases to arrive at a more definitive conclusion.

DISSEMINATED NOCARDIOSIS IDENTIFIED BY MATRIX-ASSISTED LASER DESORPTION/IONIZATION/TIME OF FLIGHT (MALDI-TOF0) MASS SPECTROMETRY

DISSEMINATED NOCARDIOSIS IDENTIFIED BY MATRIX-ASSISTED LASER DESORPTION/IONIZATION/TIME OF FLIGHT (MALDI-TOF0) MASS SPECTROMETRY

Antibiotic susceptibility of urine culture specimens in Ontario: a population-based cohort study.

Surveillance of antimicrobial resistance is essential to mitigate its impact on population health and inform local empiric treatment practices. Our aims were to evaluate urine culture specimen susceptibility from a range of diverse settings and describe antibiotic susceptibility across all organisms and compare susceptibilities to that of Escherichia coli alone. In this descriptive cohort study, we measured the prevalence of organisms in urine culture specimens using linked province-wide administrative databases. Using positive urine cultures collected in Ontario between Jan. 1, 2016, and Dec. 31, 2017, we measured susceptibility to 6 classes of antibiotics using a weighted antibiogram for all organisms compared with E. coli alone. We included 689 497 cultures derived from 569 399 patients and 879 778 test orders for specimens. For all organisms, the rates of susceptibility in the outpatient, inpatient and long-term care settings were 49.3%, 42.8% and 39.2%, respectively, for ampicillin; 83.1%, 72.7% and 69.7%, respectively, for nitrofurantoin; 80.3%, 64.8% and 73.1%, respectively, for trimethoprim-sulfamethoxazole; 87.2%, 74.1% and 66.2%, respectively, for ciprofloxacin; 90.6%, 73.6% and 85.1%, respectively, for aminoglycosides; and 82.6%, 57.5% and 73.5%, respectively, for cefazolin. We found resistance to 3 or more antibiotic classes in 20.6% of episodes for all organisms compared with 14.0% for E. coli alone. The average absolute difference in antibiotic susceptibility between all organisms and E. coli across all drugs was lowest in the outpatient setting (6.2%) and highest in the inpatient setting (14.6%). In this study, urinary organism prevalence and antimicrobial susceptibility varied across health care settings and patient populations, with implications for both antimicrobial resistance surveillance and clinical decision-making. Weighted antibiograms may be most useful for guiding empiric treatment of urinary infections in inpatient settings where the diversity of infectious organisms is higher than in the community.

Microbial diversity and antimicrobial susceptibility in endotracheal tube biofilms recovered from mechanically ventilated COVID-19 patients

In patients with acute respiratory failure, mechanical ventilation through an endotracheal tube (ET) may be required to correct hypoxemia and hypercarbia. However, biofilm formation on these ETs is a risk factor for infections in intubated patients, as the ET can act as a reservoir of microorganisms that can cause infections in the lungs. As severely ill COVID-19 patients often need to be intubated, a better knowledge of the composition of ET biofilms in this population is important. In Spring 2020, during the first wave of the COVID-19 pandemic in Europe, 31 ETs were obtained from COVID-19 patients at Ghent University Hospital (Ghent, Belgium). Biofilms were collected from the ET and the biofilm composition was determined using culture-dependent (MALDI-TOF mass spectrometry and biochemical tests) and culture-independent (16S and ITS1 rRNA amplicon sequencing) approaches. In addition, antimicrobial resistance was assessed for isolates collected via the culture-dependent approach using disc diffusion for 11 antimicrobials commonly used to treat lower respiratory tract infections. The most common microorganisms identified by the culture-dependent approach were those typically found during lung infections and included both presumed commensal and potentially pathogenic microorganisms like Staphylococcus epidermidis, Enterococcus faecalis, Pseudomonas aeruginosa and Candida albicans. More unusual organisms, such as Paracoccus yeei, were also identified, but each only in a few patients. The culture-independent approach revealed a wide variety of microbes present in the ET biofilms and showed large variation in biofilm composition between patients. Some biofilms contained a diverse set of bacteria of which many are generally considered as non-pathogenic commensals, whereas others were dominated by a single or a few pathogens. Antimicrobial resistance was widespread in the isolates, e.g. 68% and 53% of all isolates tested were resistant against meropenem and gentamicin, respectively. Different isolates from the same species recovered from the same ET biofilm often showed differences in antibiotic susceptibility. Our data suggest that ET biofilms are a potential risk factor for secondary infections in intubated COVID-19 patients, as is the case in mechanically-ventilated non-COVID-19 patients.

Fast growth can counteract antibiotic susceptibility in shaping microbial community resilience to antibiotics

Microbial communities often face external perturbations that can induce lasting changes in their composition and functions. Our understanding of how multispecies communities respond to perturbations such as antibiotics is limited, with susceptibility assays performed on individual, isolated species our primary guide in predicting community transitions. Here, we studied how bacterial growth dynamics can overcome differences in antibiotic susceptibility in determining community resilience: the recovery of the original community state following antibiotic exposure. We used an experimental community containing Corynebacterium ammoniagenes and Lactobacillus plantarum that displays two alternative stable states as a result of mutual inhibition. Although C. ammoniagenes was more susceptible to chloramphenicol in monocultures, we found that chloramphenicol exposure nonetheless led to a transition from the L. plantarum-dominated to the C. ammoniagenes-dominated community state. Combining theory and experiments, we demonstrated that growth rate differences between the two species made the L. plantarum-dominated community less resilient to several antibiotics with different mechanisms of action. Taking advantage of an observed cooperativity—a dependence on population abundance—in the growth of C. ammoniagenes, we next analyzed in silico scenarios that could compromise the high resilience of the C. ammoniagenes-dominated state. The model predicted that lowering the dispersal rate, through interacting with the growth at low population densities, could make the C. ammoniagenes state fragile against virtually any kind of antibiotic, a prediction that we confirmed experimentally. Our results highlight that species susceptibility to antibiotics is often uninformative of community resilience, as growth dynamics in the wake of antibiotic exposure can play a dominant role.

Growth and division mechanisms by which genetic resisters emerge from the rifampicin-surviving population of differentially antibiotic-susceptible mycobacterial subpopulations.

We recently reported the de novo emergence of unusually high numbers of antibiotic resisters from the in vitro cultures of Mycobacterium tuberculosis and Mycobacterium smegmatis surviving in the presence of minimum bactericidal concentration (MBC) of antituberculosis antibiotics. The resisters emerged due to multiple asymmetric divisions of elongated mother cells containing multiple nucleoids and multiple septae. We had earlier found a minor subpopulation of short-sized cells (SCs) and a major subpopulation of normal-sized cells (NCs) (10% and 90%, respectively, of the whole population), with significant difference in antibiotic susceptibility and resister generation frequency, in the in vitro cultures of M. tuberculosis, M. smegmatis, and Mycobacterium xenopi, as well as in pulmonary tuberculosis patients' sputum. However, the mechanisms of growth and division promoting the emergence of antibiotic resisters from these subpopulations remained unknown. Therefore, here, we took up the first-time study to find out the mechanism of growth and division by which antibiotic resisters emerge from the antibiotic-surviving population of the two subpopulations of M. smegmatis. M. smegmatis SCs and NCs were fractionated from mid-log phase cultures using Percoll gradient centrifugation; their purity was checked and exposed to 10×, 2×, and 0.4× MBC of rifampicin for 120 h. The colony-forming units (CFUs) were determined on rifampicin-free plates for the total population and on rifampicin-containing plates for scoring rifampicin resisters. The phenotype and the morphology of the cells at various stages of the exposure were determined using transmission electron microscopy. The dynamic growth and division mechanisms of the cells to emerge as rifampicin resisters were monitored using live-cell time-lapse imaging. The rifampicin resisters were sequenced for mutations in the rifampicin resistance determining region of rpoB gene. Statistical significance was calculated using two-tailed paired t-test, with *P ≤ 0.05 and **P ≤ 0.01. Multinucleated and multiseptated elongated cells emerged from their respective antibiotic-surviving populations. They produced a large number of sibling-daughter cells through multiple asymmetric divisions in short durations, showing abnormally high spurts in CFUs of antibiotic resisters. The CFUs were several-fold higher than that expected from the mass-doubling time of the subpopulations. Despite this commonality, the subpopulations showed specific differences in their response to different multiples of their respective MBC of rifampicin. Mycobacterial subpopulations come out of rifampicin stress by undergoing multiple nucleoid replications, multiple septation for nucleoid segregation, and acquisition of antibiotic target-specific mutations, followed by multiple asymmetric divisions to generate unusually a large number of rifampicin resisters. Because we had earlier shown that SCs and NCs are present in the pulmonary tuberculosis patients' sputum, the present findings have clinical relevance on the mechanism of emergence of antibiotic-resistant strains from mycobacterial subpopulations.

The effect of the efflux pump inhibitor Carbonyl Cyanide m-Chlorophenylhydrazone (CCCP) on the susceptibility to imipenem and cefepime in clinical strains of Acinetobacter baumannii.

IntroductionIn the last years the rapid expansion of multidrug-resistant A. baumannii strains have become a major health problem. Efflux pumps are a group of transport proteins that contribute to the development of antibiotic resistance. The aim of this study was to evaluate the effect of the efflux pump inhibitor carbonyl cyanide 3-chlorophenylhydrazone (CCCP) on the antimicrobial action of imipenem and cefepime on clinical strains of A. baumannii.Materials and methodsA total of 49 non-duplicate clinical samples were collected during January through December of 2018 from patients hospitalized in the Hospital Regional Docente de Cajamarca. Of the 49 samples obtained, the confirmatory identification of A. baumannii was performed on 47 samples by molecular methods. The amplification of the blaOXA-51-like gene was carried out by polymerase chain reaction (PCR). The determination of the minimum inhibitory concentration (MIC) was calculated using the microdilution method in culture broth. The susceptibility to both antibiotics (cefepime and imipenem) was evaluated in the presence and absence of the inhibitor carbonyl cyanide 3-chlorophenylhydrazone (CCCP).ResultsA total of 47 strains of A. baumannii were isolated: 97.87% (46/47) were resistant to Imipenem, 2.13% (1/47) of them were classified as intermediate and none of these strains were susceptible. On the other hand, 51.06% (24/47) of isolates were resistant to cefepime; 19.15% (9/47) intermediate and 29.79% (14/47) susceptible. We considered a significant difference in antibiotic susceptibility if the MIC changed at least 4 dilutions, after the addition of the inhibitor. In the case of CCCP in addition to imipenem, 2.1% (1/47) had a significant change of 4 or more reductions in MIC, 59.6% (28/47) achieved a change equal or less than 3 dilutions and 17.0% (8/47) did not have any change. In the case of CCCP with cefepime the percentage of strains with the significant change of MIC was 8.5% (4/47). On the other hand, 53.2% (24/47) presented a reduction equal or less than 3 dilutions and 12.8% (6/47) did not show changes.ConclusionIn conclusion, our results demonstrate that the use of CCCP may improve the antibiotic effect of imipenem and cefepime on clinical strains of A. baumannii. The relevance of this study is that it provides evidence that this efflux pump inhibitor may be an alternative treatment against multidrug-resistant A. baumannii.

Antibiotic resistance heterogeneity and LasR diversity within Pseudomonas aeruginosa populations from pneumonia in intensive care unit patients

This study investigated within-host heterogeneity of 66 Pseudomonas aeruginosa populations from pneumonia in 51 critically ill ventilated patients by examining 30 colonies per bronchoalveolar lavage (BAL). Differences in antibiotic susceptibility and quorum-sensing (QS) phenotypes were observed between the members of 14 (21.2%) and 10 (15.2%) populations, respectively. A significant association was found between QS deficiency and ceftazidime resistance. QS deficiency was associated with various lasR modifications, and was observed in 25 of 51 (49.0%) patients, including seven patients who received ≤48 h of ventilation. This study confirms the need to examine diverse colonies when analysing BAL cultures, particularly in β-lactam-exposed patients, to avoid missing ceftazidime- or imipenem-resistant isolates.

176. Selected Impact of the 13-valent Pneumococcal Conjugate Vaccine (PCV13) on Invasive Pneumococcal Disease (IPD) at Eight Children’s Hospitals in the United States. 2014–2019

BackgroundThe 2011 IDSA/PIDS Clinical Practice Guidelines for the Management of Community-acquired Pneumonia (CAP) in Children Older than 3 months recommended empiric treatment with either ampicillin/penicillin or ceftriaxone based on PCV13 vaccine status and the local antibiotic susceptibilities of IPD isolates. No study has addressed differences in antibiotic susceptibilities for isolates from children with pneumococcal pneumonia (PP) based on PCV13 status.MethodsInvestigators from 8 US children’s hospitals identified infants and children with IPD between 1/1/2014 and 12/31/2019. IPD was documented by positive cultures from a normally sterile site. PP diagnosis required an abnormal chest radiograph. Clinical data were recorded and isolates analyzed for serotype (ST) by the capsular swelling method and antimicrobial susceptibilities by standard methods. Administration of PCV7/13 was documented through the patient’s medical records, health care provider or a vaccine registry. Fisher Exact was performed; p< 0.05 was significant.Results690 IPD patients with isolates were available (0–18 y); 24% (166/690) of the isolates were PCV13 STs (ST3-75, ST19F-45, ST19A-36, ST7F-5, other STs-5). The most common non-PCV13 ST isolates were ST35B-60, ST23B-59, ST33F-47, ST22F-43, ST15C-35, ST23A-27, ST15B-26, ST10A-26, ST15A-23. Of non-PCV13 isolates, 41% (217/524) were among the 7 additional STs in PCV20. For children with PP (n=157), the distributions of penicillin (Table) (p=0.8) and ceftriaxone MICs were no different for isolates obtained from children regardless of prior PCV13 doses. Less than 7% of PP isolates were resistant to penicillin (MIC >2 µg/mL). Table. Minimal inhibitory concentrations (MIC) of pneumococcal isolates related to the number of PCV13 doses each patient received prior to pneumococcal pneumonia (PP).Number of PCV13 doses Penicillin MIC (µg/mL) 0 1 2 3 4 ≤ 0.125 26*1281743 0.25–0.5 42115 1 10031 2 40025 ≥4 12124*Number of pneumococcal pneumonia patients/isolates. 145/157 patients had immunization and isolate susceptibility data available for analysis.ConclusionPCV13 status should not modify empiric antibiotics for children with suspected pneumococcal CAP. 41% of non-PCV13 IPD isolates were among the 7 additional PCV20 STs.Disclosures Sheldon L. Kaplan, MD, Allergan (Research Grant or Support)Pfizer (Grant/Research Support) Tina Q. Tan, MD, Pfizer (Grant/Research Support, Other Financial or Material Support, Chair, DMSB for PCV20 vaccine) Pia S. Pannaraj, MD, MPH, AstraZeneca (Grant/Research Support)Pfizer (Grant/Research Support)Sanofi Pasteur (Advisor or Review Panel member)

Detection of Biofilm Formation of Klebsiella Pneumoniae Isolated from Medical Devices at the University Hospital of Tlemcen, Algeria

Background: Klebsiella pneumoniae is a major cause of community-acquired and nosocomial infections. This germ is responsible for acute and chronic infections, most of which are due to its ability to adhere to medical implants and form a biofilm. The objective of this work is to study the interaction between clinical isolates of K. pneumoniae and abiotic surfaces (medical devices) and some factors influencing biofilm formation. Methods: Over a period of 2 years, 115 strains of K. pneumoniae were isolated from medical devices CHU Tlemcen, most of which had a high level of resistance to cephalosporins 1st, 2nd, and 3rd generation. Their capacity to form biofilm was assessed using two 3 techniques: TCP, TP, and RCA. We determined in vitro the effects of three antimicrobial agents against planktonic and biofilm forms of K. pneumoniae. The presence of MrkD genes was detected by polymerase chain reaction (PCR). Results: According to the studied (TCP, TP, RCA) strains of K .pneumoniae isolated from urinary catheters have proved very good, forming the biofilm to those isolated from other medical devices. 24 of 115 isolated strains showed a clear difference in antibiotic susceptibility between planktonic populations and biofilm populations. They were 10-20 times higher. All strains presented a highly hydrophilic character and adhesion 2-10 times greater in PVC with respect to glass support. The MrkD gene (detected by PCR) responsible for biofilm formation was found in 22 strains of K. pneumoniae, which may explain their adhesion and therefore their pathogenicity. Conclusion: Our results show the great ability of K.pneumoniae strains to form a biofilm on medical devices, and the isolates were at least 10 times more resistant than their planktonic counterparts. In addition, we showed that the presence of type 3-encoding gene mrkD was associated with high adhesion indexes.

Progress towards a coordinated, national paediatric antimicrobial resistance surveillance programme: Staphylococcus aureus, enterococcal and Gram-negative bacteraemia in Australia.

There is increasing knowledge of antimicrobial usage in children yet limited availability of nationally representative paediatric-specific data on antimicrobial resistance. Paediatric data from this national surveillance programme are presented to explore differences between childhood and adult bloodstream infections and antimicrobial resistance surveillance. Using information collected from a prospective coordinated antimicrobial resistance surveillance programme, children ≤18 years and adults >18 years with a positive blood culture for Staphylococcus aureus, Enterococcus spp. or Gram-negative spp. presenting to one of 34 Australian hospitals during 2013-16 were evaluated. Consistent methodologies for key sepsis pathogens were employed and a comparative analysis between children and adults was conducted. There are stark contrasts between children and adults in this national antimicrobial resistance (AMR) data set. Notable differences include lower rates of AMR, different clinical and molecular phenotypes and lower mortality amongst children. The burden of Gram-negative resistance is disproportionately experienced in children, with higher odds of death with an ESBL versus non-ESBL bacteraemia in comparison with adults. These data support that children are not just 'little adults' in the AMR era, and analyses by age group are important to detect differences in antibiotic susceptibility, clinical phenotype and genetic virulence factors. Antimicrobial surveillance incorporated into routine laboratory practice is vital to inform an array of wider applications including antimicrobial guidelines, stewardship and direction for prioritization of novel antimicrobial development.

Probing the Dynamic Aspects of AcrB Function through Disulfide Bond Formation

The Resistant Nodulation Division (RND) super family member, tripartite complex AcrA-AcrB-TolC efflux pump is one of the major pumps contributing to the multidrug-resistance in Escherichia coli. AcrB is the inner membrane protein of the efflux pump that is responsible for the recognition and binding of compounds before transportation out of the cell. Understanding the dynamics of AcrB regarding the functional rotation of the monomers in a trimeric functional unit in the process of efflux of drugs is the focus of this study. For this purpose, we created 7 disulfide bond pairs in the periplasmic domain of the AcrB using site directed mutagenesis to study the rotation and flexibility of the monomers. These pairs were chosen to be in close proximity to form inter-subunit disulfide bonds. Western blot analysis revealed the formation of disulfide oligomers under non reducing condition and monomers formation under reducing condition. Furthermore, minimum inhibitory concentration assays revealed the difference in antibiotic susceptibility of single and double Cys mutation in which the majority of double Cys mutation have a higher susceptibility to drugs, supporting that disulfide bond formation has facilitate the reduction of efflux activity. This was further confirmed with EtBr accumulation assays utilizing DTT as the reducing agent. In some cases, the activity of the double Cys-mutations was partially or completely restored by DTT, confirming the deleterious effect of the corresponding disulfide bond on protein activity. These findings lead to new insight in conformational changes critical for the activity of the dynamic AcrA-AcrB-TolC efflux pump in E.coli.

Mycobacterium mageritense tattoo infection: a known complication with a novel species

Non-pigmented rapidly growing mycobacteria are nontuberculous mycobacteria (NTM) capable of producing disease. We report a case of tattoo-associated NTM infection with a novel species: Mycobacterium mageritense. A 48-year-old man presented with a two-week history of a papulopustular eruption on the shaded areas of a tattoo that had been placed five weeks prior while in the Philippines. Histopathology from punch biopsies revealed suppurative granulomatous dermatitis with acid fast bacilli present. Subsequent matrix assisted laser desorption/ionization time of flight (MALDI-TOF) mass spectrometer identified the bacteria as Mycobacterium margeritense. After consultation with infectious disease specialists and culture susceptibilities, the patient was treated with three months of dual antibiotic therapy with minocycline and moxifloxacin. The patient experienced a slow but complete resolution of clinical skin findings after the course of treatment. Since discovery in 1997, M. mageritense infection has been demonstrated in a wide spectrum of disease, predominantly skin and soft tissue infections. The species has not been previously implicated in tattoo-associated NTM infections. M. mageritense should be considered as a specific type of mycobacteria in the differential diagnosis for tattoo-associated NTM infections owing to differences in antibiotic susceptibilities compared to other NTM species.

246. Carbapenem-resistant Klebsiella (CRK) Bloodstream Infections (BSIs) Are Caused by Bacterial Populations That Are Genotypically and Phenotypically Diverse

BackgroundThe majority of bacterial BSIs are believed to stem from a single, clonal organism. We hypothesized that most CRK BSIs are caused by genetically diverse, clonal strains that exhibit different phenotypes.MethodsBlood cultures (BCs) that were positive for CRK from each of 10 patients (patients) were streaked onto agar plates, and 100 individual colonies were chosen for Illumina whole-genome sequencing. Strains from 3 patients were tested for in vitro phenotypes and virulence in mice.ResultsPatients had BSIs due to ST258 K. pneumoniae (Kp; 6), non-ST258 Kp (3), and K. michiganensis (Km; 1). 5 patients were infected with strains that differed by core genome single nucleotide polymorphism phylogeny (2–5 unique genotypes/patient) [figure]. 6 patients were infected with strains that differed by gene or plasmid content, and/or gene deletions [table]. Differences in individual patients encompassed antibiotic resistance and putative virulence genes (including mixtures of blaKPC+ and blaKPC– strains, and various capsular (CPS) and porin mutant strains). In total, BSIs in 8 of 10 patients were caused by a genotypically diverse population. In each of 3 patients, genotypically diverse ST258 Kp strains demonstrated significant differences in antibiotic susceptibility, CPS content, mucoviscosity, adherence, resistance to serum killing, and mortality rates and tissue burdens during BSIs of mice. ST258 strains from a pt with and without a KPC-bearing IncFIA plasmid differed in β-lactam susceptibility, but were equally virulent. Progressive loss of CPS in ST258 strains from another patient enhanced serum killing and adherence, and attenuated virulence. Using PCR markers to test 96 colonies per positive BC bottle, we demonstrated that strains selected by the clinical micro lab accounted for 2% to 98% of a population in different patients.ConclusionCRK causing BSIs in most patients demonstrated remarkable genotypic diversity, which impacted antibiotic susceptibility, virulence and other phenotypes. Differences were not recognized during hospitalization since clinical labs select single, morphologically distinct colonies for evaluation. Studies are needed to understand the clinical implications of our findings, diversity during other BSIs, and whether clinical lab practices need revision. Disclosures All authors: No reported disclosures.

Antibiotic resistance of Escherichia coli isolated from uncomplicated UTI in general practice patients over a 10-year period

Recommendations of first choice antibiotic therapy need to be based on actual antibiotic susceptibility data. We determined the antibiotic susceptibility of E. coli in uncomplicated UTI among women and compared the results with 2004 and 2009. In 30 sentinel general practitioner practices of Nivel Primary Care database, urine samples were collected from women with symptoms of uncomplicated UTI. Patient characteristics, E. coli susceptibility, and ESBL production were analyzed. Six hundred eighty-nine urine samples were collected; E. coli was the most isolated uropathogen (83%). Antibiotic susceptibility was stable over time except for ciprofloxacin (96% in 2004, 97% in 2009, and 94% in 2014; P < 0.05). The susceptibility to co-amoxiclav was 88%, 87%, and 92% in 2004, 2009, and 2014, respectively. The prevalence of ESBL-producing E. coli increased from 0.1% in 2004 to 2.2% in 2014 (P < 0.05). Regional differences in antibiotic susceptibility for co-trimoxazole were found being the highest in the west (88%) and the lowest in the north (72%, P = 0.021). Ciprofloxacin susceptibility was related to antibiotic use in the past 3 months (97% no use versus 90% use, P = 0.002) and age > 70 years (P = 0.005). In 2014, prescription of fosfomycin increased compared to 2009 (14.3% versus 5.6%) at the expense of co-amoxiclav, co-trimoxazole, and fluoroquinolones (P < 0.05). The susceptibility percentages to most antimicrobial agents tested were stable over 10 years’ period although the prevalence of E. coli and ESBLs significantly increased. Performance of a survey with regular intervals is warranted.

Clinical, epidemiological and microbiological characteristics of relapse and re-infection in Clostridium difficile infection

Recurrent diarrhea is a common complication of Clostridium difficile infection (CDI). Recurrent CDI (r-CDI) may be produced by the persistence of spores (relapse) or by the acquisition of a new strain (reinfection). In this study, we analyze epidemiological, clinical, microbiological and laboratory data from patients with r-CDI, relapse, and reinfection-CDI over 5 years and compared with a control group (non r-CDI). Among 60 patients with r-CDI, 36 patients had stool samples collected from two or more episodes, which were molecularly analyzed. Based on ribotyping, 63.9% of the samples were relapse, and 36.1% reinfection. In a multivariable logistic regression analysis, previous antibiotic exposure was found to be a risk factor for r-CDI (OR: 2.23; 95% CI: 1.0–4.9; p = 0.04). Patients with relapse had previous antibiotic exposure more frequently than did patients with reinfection (p = 0.03), and patients with reinfection suffered more frequently from chronic liver disease (p = 0.02) than did relapse patients. Relapse patients compared with the control group had a higher percentage of previous antibiotic exposure, although the difference was statistically no significant (73.9% vs. 91.3 p = 0.06). No significant differences for the selected variables were observed between the reinfection and control groups, although we observed a higher percentage of patients with chronic liver disease (30.8% vs 13.3%; p = 0.08). All isolates were sensitive to metronidazole and vancomycin. No significant differences in antibiotic susceptibility were found between the different groups. Sporulation and germination frequency of r-CDI were higher than non r-CDI (p = 0.02 and p < 0.01, respectively). Nevertheless, there were statistically not significant differences between the relapse and reinfection groups. Both frequencies were compared between the first and second episode of CDI for the relapse and reinfection groups, but differences were not observed to be statistically significant. In conclusion, our study showed that the recurrence of CDI was associated with antibiotic use and sporulation/germination frequency, regardless of relapse or reinfection. The use of antibiotics would produce a dysbiosis and favor the persistence of the C. difficile spores and relapse. A possible alteration of the intestinal microbiota and the bile salts produced by chronic liver disease could favor reinfection.

Antimicrobial susceptibility, risk factors and prevalence of bla cefotaximase, temoneira, and sulfhydryl variable genes among Escherichia coli in community-acquired pediatric urinary tract infection.

INTRODUCTION:The emergence of extended-spectrum beta-lactamase (ESBL)-producing Escherichia coli has become an important challenge among pediatric patients with community-acquired urinary tract infection (UTI).OBJECTIVES:The aim of this study was to assess the antimicrobial susceptibility patterns, associated risk factors and to survey the frequency of bla cefotaximase (CTX-M), bla temoneira (TEM), and bla sulfhydryl variable (SHV) genotypes in ESBL-producing E. coli isolated from children with community-acquired UTI.METHODS:This was a prospective study conducted from November 2012 to March 2016 in a tertiary care center. E. coli isolated in urine cultures from children aged ≤18 years was identified and confirmed for ESBL production. ESBL-positive strains were screened for ESBL encoding genes. Chi-square test and Fisher's exact test were used to compare the difference in antibiotic susceptibility with respect to ESBL positive and negative, and binary logistic regression was used to identify the risk factors associated with ESBL production.RESULTS:Among 523 E. coli isolates, 196 (37.5%) were ESBL positive, >90% were resistant to cephalosporins, and 56% were resistant to fluoroquinolones. Least resistance was observed for imipenem, netilmicin, and nitrofurantoin (2%, 8.6%, 15.3%). Association between ESBL production and drug resistance was significant for ceftazidime (P < 0.001), cefixime (P < 0.001), cefotaxime (P = 0.010), ceftazidime-clavulanic acid (P < 0.001), levofloxacin (P = 0.037), and gentamicin (P = 0.047) compared to non-ESBL E. coli. CTX-M gene was the most prevalent (87.5%), followed by TEM (68.4%) and SHV (3.1%). Previous history of UTI and intake of antibiotics were the common risk factors.CONCLUSION:ESBL-producing E. coli from community-acquired pediatric UTI carries more than one type of beta-lactamase coding genes correlating their increased antibiotic resistance. Aggressive infection control policy, routine screening for detecting ESBL isolates in clinical samples, and antimicrobial stewardship are the keys to prevent their dissemination in community settings.