Concern levels of beta-lactamase-producing Gram-negative bacilli in hospital wastewater: hotspot of antimicrobial resistance in Latin-America.

Hospital wastewater (HWW) discharges are among the main sources of antibiotic-resistant bacteria. This study detected a high frequency of beta-lactamase-producing Gram-negative Bacilli in HWW of different geographical regions of Colombia, even in the presence of the wastewater treatment plant, highlighting the importance of regulating these environments in developing countries.

The remarkable genetic relationship between Staphylococcus aureus isolates from hemodialysis patients and their household contacts: Homes as an important source of colonization and dissemination.

INTRODUCTION: Staphylococcus aureus is a successful pathogen in hospital and community. Hemodialysis patients have high colonization rates. Interactions between them and their household contacts, are an opportunity to understand the S. aureus colonization between hospitals and community. This study aims to determine the clinical and epidemiological characteristics of S. aureus colonization in hemodialysis patients and their household contacts, as well as the genetic relationship between their isolates. METHODS: A cross-sectional study was conducted on hemodialysis patients from hospital-associated dialysis center in Medellín-Colombia, and their household contacts between 2019 and 2020. Colonization was assessed in the nostrils for household contacts and nostrils and skin around the catheter insertion for hemodialysis patients. Epidemiological information was obtained, and colonization was evaluated in their pets' oral cavities. Bacterial identification and susceptibility were assessed using phenotypic and molecular methods. Molecular typing included SCCmec typing, pulsed-field gel electrophoresis (PFGE), spa typing, and virulence factor detection. RESULTS: Colonization frequency was 35.6% (n = 16/45) in patients (87.5% MSSA- 12.5% MRSA) and 43.1% (n = 53/123) in household contacts (88.7% MSSA-11.3% MRSA). Of 45 homes, 77.8% presented colonized people. Colonization was detected in at least two household members in 46.7% of homes, of which 52.4% had a genetic relationship. Colonization was 16% (n = 4/25) in pets (75% MRSA-25% MSSA). The most frequent clonal complex was CC8 (15.6%), and the spa typing revealed high diversity. CONCLUSION: This study shows a high frequency of colonization by S. aureus in both hemodialysis patients and their household contacts and a significant genetic relationship between their isolates. This demonstrates an exchange of this bacterium and that homes are an important source of colonization to patients, highlighting the need for prevention strategies outside the hospital to avoid future infections, and the importance of the populations with permanent transit between the two environments.

Climatological and Epidemiological Conditions Are Important Factors Related to the Abundance of blaKPC and Other Antibiotic Resistance Genes (ARGs) in Wastewater Treatment Plants and Their Effluents, in an Endemic Country.

Several physicochemical and season factors have been related to the abundance of antibiotic resistance genes (ARGs) in wastewater treatment plants (WWTPs), considered hotspots of bacterial resistance. However, few studies on the subject have been carried out in tropical countries endemic for resistance mechanisms such as blaKPC. In this study, the occurrence of ARGs, particularly blaKPC, was determined throughout a WWTP, and the factors related to their abundance were explored. In 2017, wastewater samples were taken from a WWTP in Colombia every 15 days for 6 months, and a total of 44 samples were analyzed by quantitative real-time PCR. sul1, sul2, blaKPC, and ermB were found to be the most prevalent ARGs. A low average reduction of the absolute abundance ARGs in effluent with respect to influent was observed, as well as a greater absolute abundance of ARGs in the WWTP effluent in the rainy season. Factors such as temperature, pH, oxygen, total organic carbon (TOC), chemical oxygen demand (COD), and precipitation were significantly correlated with the absolute abundance of several of the ARGs evaluated. A generalized linear mixed-effects model analysis showed that dissolved oxygen and precipitation in the sampling day were important factors related to the absolute concentration of blaKPC over time. In conclusion, the abundance of ARGs in the WWTP could be influenced by endemic conditions and physicochemical and climatological parameters. Therefore, it is necessary to continuously monitor clinical relevant genes in WWTPs from different global regions, even more so in low-income countries where sewage treatment is limited.

Irreversible inactivation of carbapenem-resistant Klebsiella pneumoniae and its genes in water by photo-electro-oxidation and photo-electro-Fenton - Processes action modes.

Carbapenem-resistant Klebsiella pneumoniae is a critical priority pathogen according to the World Health Organization's classification. Effluents of municipal wastewater treatment plants (EWWTP) may be a route for K. pneumoniae dissemination. Herein, the inactivation of this microorganism in simulated EWWTP by the photo-electro-oxidation (PEO) and photo-electro-Fenton (PEF) processes was evaluated. Firstly, the disinfecting ability and action pathways of these processes were established. PEO achieved faster K. pneumoniae inactivation (6 log units in 75 min of treatment) than the PEF process (6 log units in 105 min of treatment). PEO completely inactivated K. pneumoniae due to the simultaneous action of UVA light, electrogenerated H2O2, and anodic oxidation pathways. The slower inactivation of K. pneumoniae when using PEF was related to interfering screen effects of iron oxides on light penetration and the diffusion of the bacteria to the anode. However, both PEO and PEF avoided the recovery and regrowth of treated bacteria (with no detectable increase in the bacteria concentration after 24 h of incubation). In addition to the bacteria evolution, the effect of treatment processes on the resistance gene was examined. Despite inactivation of K. pneumoniae by PEF was slower than by PEO, the former process induced a stronger degrading action on the gene, conferring the resistance to carbapenems (PEF had a Ct value of 24.92 cycles after 105 min of treatment, while PEO presented a Ct of 19.97 cycles after 75 min). The results of this research indicate that electrochemical processes such as PEO and PEF are highly effective at dealing with resistant K. pneumoniae in the EWWTP matrix.

Metagenomic analysis of urban wastewater resistome and mobilome: A support for antimicrobial resistance surveillance in an endemic country.

In developing countries, where high levels of antimicrobial resistance are observed in hospitals, the surveillance of this phenomenon in wastewater treatment plants (WWTPs) and the environment is very limited, especially using cutting-edge culture-independent methods. In this study, the composition of bacterial communities, the resistome and mobilome (the pool of antibiotic resistance genes (ARGs) and mobile genetic elements (MGEs), respectively) at a WWTP were determined using shotgun metagenomics and culture-based approaches. Wastewater samples were collected at four sampling points of a WWTP in Antioquia, Colombia. A total of 24 metagenomes were analyzed. Specifically, there were marked differences in bacterial community composition, resistome, and mobilome, according to the WWTP sampling points. Bacterial families of clinical importance such as Moraxellaceae, Aeromonadaceae, and Enterobacteriaceae were mainly detected in the WWTP influent and effluent samples. Genes encoding resistance to macrolide-lincosamide-streptogramin, ß-lactams, and those conferring multidrug resistance (e.g., acrB, adeG, and mexD) were the most abundant. Moreover, some clinically important ARGs such as blaKPC-2 and blaCTX-M, and others not reported locally, such as blaTEM-196, blaGES-23, blaOXA-10, mcr-3, and mcr-5 were frequently detected. Co-occurrence network analyses indicated a significant association of ARGs such as blaOXA-58 and blaKPC genes with Aeromonadaceae and Enterobacteriaceae. Among the markers of MGEs, intI1 and ISCR8 were the most frequently detected. Altogether, this work reveals the importance of shotgun metagenomics and culture-based approaches in antimicrobial resistance studies. The findings also support that WWTPs are hotspots for antimicrobial resistance, whose analysis constitutes a powerful tool to predict the impact of antimicrobial resistance in a population.

Staphylococcus aureus colonization increases the risk of bacteremia in hemodialysis patients: a molecular epidemiology approach with time-dependent analysis.

BACKGROUND: Bacteremia is the second cause of death in hemodialysis patients and colonization may be a risk factor. We analyzed the association between Staphylococcus aureus or multidrug-resistant Gram-negative bacteria colonization and bacteremia in hemodialysis patients. METHODS: A prospective cohort study was conducted. Colonization status was determined at baseline, 2, and 6 months later. The time-to-first-bacteremia was analyzed using the baseline status and time-dependent nature of colonization. The recurrence of bacteremia given colonization status was evaluated using a Poisson regression model. The genetic relatedness between isolates that colonized and caused bacteremia were established by molecular typing methods. RESULTS: Seventy-one patients developed bacteremia over the course of follow-up, with the majority of cases being caused by S aureus (n = 28; 39.4%) and only three caused by multidrug-resistant Gram-negative bacteria. S aureus colonization was associated with an increased risk of bacteremia in time-dependent analysis (HR:4.64; 95%CI: 1.72-12.53) and with recurrence of infection in Poisson model (IRR:5.90, 95%CI: 2.29-15.16). Molecular methods revealed that 77.8% of patients with S aureus bacteremia were colonized with the same strain that caused the infection. CONCLUSIONS: S aureus is a cause of endogenous infection in hemodialysis patients. Colonization is associated with both time-to-first-bacteremia and the recurrence of infection. The prompt identification of colonized patients and the evaluation of decolonization protocols are needed.

A longitudinal study shows intermittent colonization by Staphylococcus aureus with a high genetic diversity in hemodialysis patients.

Staphylococcus aureus colonization increases the risk of invasive infections in different groups of patients. We analyzed the dynamics and factors associated with S. aureus colonization in hemodialysis patients. A longitudinal study was conducted at a dialysis center associated with a tertiary health care institution. S. aureus colonization was assessed three times in nostrils and on the skin and was classified as absent, intermittent or persistent. The molecular analysis included pulsed-field gel electrophoresis (PFGE) and spa-typing. Clonal complex was inferred from spa-typing. A model of generalized estimating equations was performed to determine the factors associated with colonization. A total of 210 patients were included. Colonization by methicillin-susceptible (MSSA) and methicillin-resistant (MRSA) isolates was 29.1 % vs. 4.8 %, 29.2 % vs. 6.7 % and 24.1 % vs. 7.1 % in the first, second and third screenings respectively. Most of the colonized patients were intermittent carriers (77.8 %, n = 63). PFGE and spa-typing revealed a high genetic diversity. One third (33.3 %) of the carriers classified as persistent had different clones during follow-up. Clonal complex 8 was frequent among MSSA (28 %) and MRSA (59 %) isolates. Current smoking (OR:7.22, 95 %CI 2.24-23.27), Charlson index (OR:1.22, 95 %CI 1.03-1.43) and previous infection by S. aureus (OR:2.41; 95 %CI:1.09-5.30) were associated with colonization by this microorganism. Colonization increased the risk of bacteremia (HR = 4.9; 95 % CI: 1.9-12.9). In conclusion, the colonization by S. aureus in hemodialysis patients changes over time and acquisition of new clones is a frequent event. These results evidence that patients are repeatedly recolonizing from hospitals, dialysis units and their homes. On the other hand, factors not associated with healthcare, as smoking, can increase the risk of colonization.

High frequency of colonization by diverse clones of beta-lactam-resistant Gram-negative bacilli in haemodialysis: different sources of transmission outside the renal unit?

Introduction. While colonization by Staphylococcus aureus in haemodialysis patients has been assessed, knowledge about colonization by beta-lactam-resistant Gram-negative bacilli is still limited.Aim. To describe clinical and molecular characteristics in haemodialysis patients colonized by S. aureus (MSSA-MRSA) and beta-lactam-resistant Gram-negative bacilli in an ambulatory renal unit.Methodology. The study included patients with central venous catheters in an outpatient haemodialysis facility in Medellín, Colombia (October 2017-October 2018). Swab specimens were collected from the nostrils and skin around vascular access to assess colonization by S. aureus (MSSA-MRSA). Stool samples were collected from each patient to evaluate beta-lactam-resistant Gram-negative bacilli colonization. Molecular typing included PFGE, multilocus sequence typing (MLST), spa typing and enterobacterial repetitive intergenic consensus-PCR (ERIC). Clinical information was obtained from medical records and personal interview.Results. A total of 210 patients were included in the study. S. aureus colonization was observed in 33.8 % (n=71) of the patients, 4.8 % (n=10) of which were colonized by methicillin-resistant S. aureus. Stool samples were collected from 165 patients and of these 41.2 % (n=68) and 11.5 % (n=19) were colonized by extended-spectrum-beta-lactamase-producing (ESBL) and carbapenem-resistant bacilli, respectively. Typing methods revealed high genetic diversity among S. aureus and ESBL-producing Gram-negative bacilli (ESBL-GNB). Antibiotic use and hospitalization in the previous 6 months were observed in more than half of the studied population.Conclusion. The high colonization by ESBL-GNB in haemodialysis patients shows evidence for the need for stronger surveillance, not only for S. aureus but also for multidrug-resistant bacilli in order to avoid their spread. Additionally, the high genetic diversity suggests other sources of transmission outside the renal unit instead of horizontal transmission between patients.

Multidrug resistance and diversity of resistance profiles in carbapenem-resistant Gram-negative bacilli throughout a wastewater treatment plant in Colombia.

OBJECTIVES: Carbapenem-resistant Gram-negative bacilli (CRGNB) have been reported in different wastewater treatment plants (WWTPs) throughout the world; however, few studies have described the antimicrobial resistance profile in different CRGNB throughout WWTPs, information that would identify points of selection of resistant bacteria. The objective of this work was to characterize the resistance profile of CRGNB harbouring blaKPC-2 from a Colombian WWTP. METHODS: Six samples were taken from four points of a WWTP. CRGNB were selected in chromID® CARBA and identified by 16S rRNA. Carbapenemases were determined by polymerase chain reaction (PCR), and susceptibility was assessed using VITEK2. RESULTS: One hundred and forty-two CRGNB harbouring blaKPC-2 were detected: 41% corresponded to Aeromonas spp. (n = 58) and 59% to Enterobacteriaceae. To establish the resistance profile, 50% of the isolates were selected proportionally by family and sampling point (26 Aeromonadaceae and 45 Enterobacteriaceae). All Enterobacteriaceae showed resistance to carbapenems and penicillins + inhibitors, high percentages of resistance to ceftriaxone (88.9%), and ciprofloxacin (44.4%), and low resistance to other antibiotics (>30%). In Aeromonadaceae, 76.9% were resistant to ceftriaxone, 58% to carbapenems, and 65.4% to ciprofloxacin. Twenty-one resistance profiles were observed, the most common of which were resistant to penicillins + inhibitor, cephalosporins (third to fourth generation), and carbapenems (19%). The percentage of multidrug resistance was 91% and was similar at all points of the WWTP. CONCLUSIONS: The high frequency of multidrug resistance and great diversity of resistance profiles observed throughout the WWTP is of concern, and shows the role of WWTP as a reservoir and dissemination source of antimicrobial resistance to water sources.

Risk factors and survival of patients infected with carbapenem-resistant Klebsiella pneumoniae in a KPC endemic setting: a case-control and cohort study.

BACKGROUND: Many gaps in the burden of resistant pathogens exist in endemic areas of low- and middle-income economies, especially those endemic for carbapenem resistance. The aim of this study is to evaluate risk factors for carbapenem-resistance, to estimate the association between carbapenem-resistance and all-cause 30-day mortality and to examine whether mortality is mediated by inappropriate therapy. METHODS: A case-control and a cohort study were conducted in one tertiary-care hospital in Medellín, Colombia from 2014 to 2015. Phenotypic and genotypic characterization of isolates was performed. In the case-control study, cases were defined as patients infected with carbapenem-resistant K. pneumoniae (CRKP) and controls as patients infected with carbapenem-susceptible K. pneumoniae (CSKP). A risk factor analysis was conducted using logistic regression models. In the cohort study, the exposed group was defined as patients infected with CRKP and the non-exposed group as patients infected with CSKP. A survival analysis using an accelerated failure time model with a lognormal distribution was performed to estimate the association between carbapenem resistance and all-cause 30-day-mortality and to examine whether mortality is mediated by inappropriate therapy. RESULTS: A total of 338 patients were enrolled; 49 were infected with CRKP and 289 with CSKP. Among CRKP isolates CG258 (n = 29), ST25 (n = 5) and ST307 (n = 4) were detected. Of importance, every day of meropenem (OR 1.18, 95%CI 1.10-1.28) and cefepime (OR 1.22, 95%CI 1.03-1.49) use increase the risk of carbapenem resistance. Additional risk factors were previous use of ciprofloxacin (OR 2.37, 95%CI 1.00-5.35) and urinary catheter (OR 2.60, 95%CI 1.25-5.37). Furthermore, a significant lower survival time was estimated for patients infected with CRKP compared to CSKP (Relative Times 0.44, 95%CI 0.24-0.82). The strength of association was reduced when appropriate therapy was included in the model (RT = 0.81 95%CI 0.48-1.37). CONCLUSION: Short antibiotic courses had the potential to reduce the selection and transmission of CRKP. A high burden in mortality occurred in patients infected with CRKP in a KPC endemic setting and CRKP leads to increased mortality via inappropriate antibiotic treatment. Furthermore, dissemination of recognized hypervirulent clones could add to the list of challenges for antibiotic resistance control.

Inactivation of carbapenem-resistant Klebsiella pneumoniae by photo-Fenton: Residual effect, gene evolution and modifications with citric acid and persulfate.

The photo-Fenton process application to eliminate carbapenem-resistant Klebsiella pneumoniae, an antibiotic-resistant priority pathogen, was evaluated. Initially, reagents concentration effect was tested and under suitable conditions (5 mg L-1 of Fe2+ and 50 mg L-1 of H2O2) complete bacteria inactivation by action of hydroxyl radical and UVA plus hydrogen peroxide was achieved at 120 min. The process presented a strong residual disinfecting effect when light was turned off at only 20 min. Besides, the cultivability of treated K. pneumoniae in a selective medium containing carbapenem antibiotics was considered. bla-KPC, gene responsible for the resistance, evolution was also assessed. The bacteria response to carbapenem antibiotics was higher as the treatment time increased. In turn, bla-KPC gene remained when K. pneumoniae was completely inactivated (120 min); nevertheless, treatment times longer than 120 min diminished bla-KPC presence. Finally, the photo-Fenton process and its modifications (citric acid addition or persulfate anion instead hydrogen peroxide) were applied to a real hospital wastewater in Colombia. In such complex matrix, the conventional photo-Fenton system reached a moderate disinfection (∼3.5 log-units at 300 min). Meanwhile, in presence of citric acid total inactivation was completed at the same time. Interestingly, the H2O2 substitution by persulfate strongly accelerated the microorganism elimination, achieving the 6-log-units reduction after only 60 min of process action. Thus, the effective elimination of K. pneumoniae from water by the modified photo-Fenton evidenced the potential applicability of this process to limit the proliferation of antibiotic resistant bacteria.

Detection of carbapenem resistance genes in Pseudomonas aeruginosa isolates with several phenotypic susceptibility profiles / Detección de genes de resistencia a carbapenémicos en aislados de Pseudomonas aeruginosa con diferentes perfiles de susceptibilidad fenotípica

Abstract Introduction: Pseudomonas aeruginosa display several resistance mechanisms to carbapenems and such variety makes it difficult to infer from the antibiogram. The aim of this study was to determine the carbapenem resistance genes in P. aeruginosa isolates with different profiles of phe-notypic susceptibility to these antibiotics. Materials and methods: From a microbial collection of P aeruginosa isolates from infected patients, 40 isolates with different carbapenem resistance profiles were selected. The carbapenemases genes, and expression of the OprD porin, the MexAB-OprM efflux pump and the p-lactamase AmpC were determined. Results: From a total of 40 isolates evaluted, in 21 (52.5%) any mechanism of resistance evaluated were detected. In the meropenem-resistant group, overexpression of AmpC (n = 1) and decreased expression of MexAB-OprM (n = 2) and OprD (n = 1) were found. A decrease in the expression of MexAB-OprM was observed in imipenem-resistant group (n = 3) and mutations in the gene encoding the OprD porin (n = 1). Finally, the presence of carbapenemases (VIM, n= 3, KPC-2 / VIM, n = 1) was detected in imipenem-meropenem resistant isolates. Conclusion: The phenotypic susceptibility profiles in P aeruginosa isolates were not explained by the molecular mechanisms explored, with the exception of carbapenemase-producing isolates. These results evidence the complexity of the antibiotic resistance mechanisms involved in this bacterium.

Resumen Introducción: Pseudomonas aeruginosa presenta diferentes mecanismos de resistencia a los carbapenémicos, dificultando su inferencia a partir del antibiograma. El objetivo fue determinar los genes de resistencia a car-bapenémicos en aislados de Pseudomonas aeruginosa con diferentes perfiles de susceptibilidad a estos antibióticos. Materiales y métodos: A partir de una colección microbiana de aislados de P. aeruginosa provenientes de pacientes infectados se seleccionaron 40 aislados con diferentes perfiles de resistencia a carbapenémicos y en los cuales se determinaron los genes de car-bapenemasas, la expresión de la porina OprD, la bomba de expulsión MexAB-OprM y la betalactamasa AmpC. Resultados: El 52,5 % de los aislados no presentó ninguno de los mecanismos de resistencia evaluados. En los resistentes a meropenem se encontró sobreexpresión de AmpC (n=1) y disminución de la expresión de MexAB-OprM (n=2) y OprD (n=1). En los resistentes a imipenem se observó disminución en la expresión de MexAB-OprM (n=3) y mutaciones en el gen que codifica la porina OprD (n=1). En aislados resistentes a imipenem y meropenem se detectó la presencia de carbapenemasas (VIM, n=3, KPC/VIM, n=1). Conclusión: Los mecanismos moleculares hallados no explican el fenotipo de resistencia a carbapenémicos, excepto en los aislados productores de carbapenemasas. Estos resultados evidencian la complejidad de los mecanismos implicados en la resistencia antibiótica en esta bacteria.

Colistin Resistance in Carbapenem-Resistant Klebsiella pneumoniae Mediated by Chromosomal Integration of Plasmid DNA.

Here we describe the spread of colistin resistance in clinical isolates of carbapenem-resistant Klebsiella pneumoniae in Medellín, Colombia. Among 32 isolates collected between 2012 and 2014, 24 showed genetic alterations in mgrB Nineteen isolates belonged to sequence type 512 (ST512) (or its single locus variant [SLV]) and harbored an 8.1-kb hsdMSR insertion corresponding to ISKpn25, indicating a clonal expansion of the resistant strain. The insertion region showed 100% identity to several plasmids, suggesting that the colistin resistance is mediated by chromosomal integration of plasmid DNA.

Molecular epidemiology of carbapenem resistant gram-negative bacilli from infected pediatric population in tertiary - care hospitals in Medellín, Colombia: an increasing problem.

BACKGROUND: Gram-negative bacilli are a cause of serious infections in the pediatric population. Carbapenem are the treatment of choice for infections caused by multidrug-resistant Gram-negative bacilli, but the emergence of carbapenem resistance has substantially reduced access to effective antimicrobial regimens. Children are a population vulnerable to bacterial infections and the emergence of resistance can worsen prognosis. The aim of this study is to describe the clinical and molecular characteristics of infections caused by carbapenem-resistant Gram-negative bacilli in pediatric patients from five tertiary-care hospitals in Medellín, Colombia. METHODS: A cross-sectional study was conducted in five tertiary-care hospitals from June 2012 to June 2014. All pediatric patients infected by carbapenem-resistant Gram-negative bacilli were included. Clinical information for each patient was obtained from medical records. Molecular analyses included PCR for detection of bla VIM, bla IMP bla NDM, bla OXA-48 and bla KPC genes and PFGE and MLST for molecular typing. RESULTS: A total of 59 patients were enrolled, most of them less than 1 year old (40.7 % n = 24), with a previous history of antibiotic use (94.9 %; n = 56) and healthcare-associated infections - predominately urinary tract infections (31.0 %; n = 18). Klebsiella pneumoniae was the most frequent bacteria (47.4 %), followed by Enterobacter cloacae (40.7 %) and Pseudomonas aeruginosa (11.9 %). For K. pneumoniae, KPC was the predominant resistance mechanism (85.7 %; n = 24) and ST14 was the most common clone (39.3 % n = 11), which included strains closely related by PFGE. In contrast, E. cloacae and P. aeruginosa were prevailing non-carbapenemase-producing isolates (only KPC and VIM were detected in 1 and 3 isolates, respectively) and high genetic diversity according to PFGE and MLST was found in the majority of the cases. CONCLUSIONS: In recent years, increasing carbapenem-resistant bacilli in children has become in a matter of great concern. It is important to conduct systemic surveillance and take measures to prevent dissemination of multidrug-resistant bacteria.

A Two-Year Surveillance in Five Colombian Tertiary Care Hospitals Reveals High Frequency of Non-CG258 Clones of Carbapenem-Resistant Klebsiella pneumoniae with Distinct Clinical Characteristics.

The global spread of carbapenem-resistant Klebsiella pneumoniae (CR-Kp) has been largely associated with sequence type 258 (ST258) and its related variants (clonal group 258 [CG258]). Here we describe the molecular epidemiology of CR-Kp from five tertiary care hospitals in Medellín, the second largest city in Colombia. All CR-Kp-infected patients admitted from June 2012 to June 2014 were included (n = 193). Patients' clinical information was obtained from medical records. Carbapenemase KPC, VIM, IMP, NDM, and OXA-48 genes were detected by PCR. A CG258-tonB79 cluster-specific real-time PCR (targeting the multilocus sequence type [MLST] tonB79 allele), pulsed-field gel electrophoresis (PFGE), and MLST analysis were performed for typing. Remarkably, 62.2% (n = 120) of isolates were from STs unrelated to CG258 (non-CG258). KPC-3 predominated in CG258 isolates (86.3%), while KPC-2 prevailed in non-CG258 isolates (75.5%) (P < 0.001). Multidrug resistance (MDR) frequency was significantly higher in CG258 strains (91.4% versus 56.1%; P < 0.001). ST512 (a single-locus variant of ST258) is the main ST in CG258 (96.3%), and isolates in this group showed closely related pulsotype and similar resistance gene profiles, suggesting the clonal spread of this strain. In contrast, high heterogeneity of STs (34/54), including eight novel STs, was found in non-CG258 isolates. Among non-CG258 isolates, ST14 (13.3%; n = 16) and ST307 (14.2%; n = 17) were the most frequent, and they showed distinct molecular and clinical characteristics in comparison to CG258 isolates. Our results suggest that the dissemination of carbapenem resistance in Medellín is due to heterogeneous K. pneumoniae clones, likely the result of horizontal transmission of KPC in different unrelated lineages, further highlighting the challenge in CR-Kp infection control and the need for a multifocal intervention.

Similar frequencies of Pseudomonas aeruginosa isolates producing KPC and VIM carbapenemases in diverse genetic clones at tertiary-care hospitals in Medellín, Colombia.

Carbapenem-resistant Pseudomonas aeruginosa has become a serious health threat worldwide due to the limited options available for its treatment. Understanding its epidemiology contributes to the control of antibiotic resistance. The aim of this study was to describe the clinical and molecular characteristics of infections caused by carbapenem-resistant P. aeruginosa isolates in five tertiary-care hospitals in Medellín, Colombia. A cross-sectional study was conducted in five tertiary-care hospitals from June 2012 to March 2014. All hospitalized patients infected by carbapenem-resistant P. aeruginosa were included. Clinical information was obtained from medical records. Molecular analyses included PCR for detection of bla(VIM), bla(IMP), bla(NDM), bla(OXA-48), and bla(KPC) genes plus pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing (MLST) for molecular typing. A total of 235 patients were enrolled: 91.1% of them were adults (n = 214), 88.1% (n = 207) had prior antibiotic use, and 14.9% (n = 35) had urinary tract infections. The bla(VIM-2) and bla(KPC-2) genes were detected in 13.6% (n = 32) and 11.5% (n = 27), respectively, of all isolates. Two isolates harbored both genes simultaneously. For KPC-producing isolates, PFGE revealed closely related strains within each hospital, and sequence types (STs) ST362 and ST235 and two new STs were found by MLST. With PFGE, VIM-producing isolates appeared highly diverse, and MLST revealed ST111 in four hospitals and five new STs. These results show that KPC-producing P. aeruginosa is currently disseminating rapidly and occurring at a frequency similar to that of VIM-producing P. aeruginosa isolates (approximately 1:1 ratio) in Medellín, Colombia. Diverse genetic backgrounds among resistant strains suggest an excessive antibiotic pressure resulting in the selection of resistant strains.

Differences in epidemiological and molecular characteristics of nasal colonization with Staphylococcus aureus (MSSA-MRSA) in children from a university hospital and day care centers.

BACKGROUND: Clinical significance of Staphylococcus aureus colonization has been demonstrated in hospital settings; however, studies in the community have shown contrasting results regarding the relevance of colonization in infection by community-associated MRSA (CA-MRSA). In Colombia there are few studies on S. aureus colonization. The aim of this study was to determine the molecular and epidemiological characteristics of nasal colonization by S. aureus (MSSA-MRSA) in children from a university hospital and day care centers (DCCs) of Medellin, Colombia. METHODS: An observational cross-sectional study was conducted in 400 children (200 in each setting), aged 0 months to 5 years, during 2011. Samples were collected from each nostril and epidemiological information was obtained from the parents. Genotypic analysis included spa typing, PFGE, MLST, SCCmec typing, detection of genes for virulence factors and agr groups. RESULTS: Frequency of S. aureus colonization was 39.8% (n = 159) (hospital 44.5% and DCCs 35.0%) and by MRSA, 5.3% (n = 21) (hospital 7.0% and DCCs 3.5%). Most S. aureus colonized children were older than two years (p = 0.005), the majority of them boys (59.1%), shared a bedroom with a large number of people (p = 0.028), with history of ß-Lactamase inhibitors usage (p = 0.020). MSSA strains presented the greatest genotypic diversity with 15 clonal complexes (CC). MRSA isolates presented 6 CC, most of them (47.6%) belonged to CC8-SCCmec IVc and were genetically related to previously reported infectious MRSA strains. CONCLUSION: Differences in epidemiological and molecular characteristics between populations may be useful for the understanding of S. aureus nasal colonization dynamics and for the design of strategies to prevent S. aureus infection and dissemination. The finding of colonizing MRSA with similar molecular characteristics of those causing infection demonstrates the dissemination capacity of S. aureus and the risk of infection among the child population.

A comparison of methicillin-resistant and methicillin-susceptible Staphylococcus aureus reveals no clinical and epidemiological but molecular differences.

Most studies on Staphylococcus aureus have focused on the molecular epidemiology of methicillin-resistant S. aureus (MRSA) infections. In contrast, little information is available regarding the molecular epidemiology of currently circulating methicillin-susceptible S. aureus (MSSA) isolates in hospital settings, an epoch when the epidemiology of S. aureus has undergone significant changes. We conducted a cross-sectional study to compare the clinical, epidemiological, and genetic characteristics of MSSA and MRSA isolates at 3 tertiary-care hospitals in Medellín, Colombia, from February 2008 to June 2010. The infections were classified according to the Centers for Disease Control and Prevention (CDC) definitions. Genotypic analysis included spa typing, multilocus sequence typing (MLST) and staphylococcal cassette chromosome (mec) (SCCmec) typing. A total of 810 patients was enrolled. One hundred infections (12.3%) were classified as community-associated (31 CA-MSSA, 69 CA-MRSA), 379 (46.8%) as healthcare-associated community-onset (136 HACO-MSSA, 243 HACO-MRSA), and 331 (40.9%) as healthcare-associated hospital-onset (104 HAHO-MSSA, 227 HAHO-MRSA). Genotype analyses showed a higher diversity and a more varied spa type repertoire in MSSA than in MRSA strains. Most of the clinical-epidemiological characteristics and risk factors evaluated did not allow for discriminating MRSA- from MSSA-infected patients. The lack of equivalence among the genetic backgrounds of the major MSSA and MRSA clones would suggest that the MRSA clones are imported instead of arising from successful MSSA clones. This study emphasizes the importance of local surveillance to create public awareness on the changing S. aureus epidemiology.

CC8 MRSA strains harboring SCCmec type IVc are predominant in Colombian hospitals.

BACKGROUND: Recent reports highlight the incursion of community-associated MRSA within healthcare settings. However, knowledge of this phenomenon remains limited in Latin America. The aim of this study was to evaluate the molecular epidemiology of MRSA in three tertiary-care hospitals in Medellín, Colombia. METHODS: An observational cross-sectional study was conducted from 2008-2010. MRSA infections were classified as either community-associated (CA-MRSA) or healthcare-associated (HA-MRSA), with HA-MRSA further classified as hospital-onset (HAHO-MRSA) or community-onset (HACO-MRSA) according to standard epidemiological definitions established by the U.S. Centers for Disease Control and Prevention (CDC). Genotypic analysis included SCCmec typing, spa typing, PFGE and MLST. RESULTS: Out of 538 total MRSA isolates, 68 (12.6%) were defined as CA-MRSA, 243 (45.2%) as HACO-MRSA and 227 (42.2%) as HAHO-MRSA. The majority harbored SCCmec type IVc (306, 58.7%), followed by SCCmec type I (174, 33.4%). The prevalence of type IVc among CA-, HACO- and HAHO-MRSA isolates was 92.4%, 65.1% and 43.6%, respectively. From 2008 to 2010, the prevalence of type IVc-bearing strains increased significantly, from 50.0% to 68.2% (p = 0.004). Strains harboring SCCmec IVc were mainly associated with spa types t1610, t008 and t024 (MLST clonal complex 8), while PFGE confirmed that the t008 and t1610 strains were closely related to the USA300-0114 CA-MRSA clone. Notably, strains belonging to these three spa types exhibited high levels of tetracycline resistance (45.9%). CONCLUSION: CC8 MRSA strains harboring SCCmec type IVc are becoming predominant in Medellín hospitals, displacing previously reported CC5 HA-MRSA clones. Based on shared characteristics including SCCmec IVc, absence of the ACME element and tetracycline resistance, the USA300-related isolates in this study are most likely related to USA300-LV, the recently-described 'Latin American variant' of USA300.