Caracterización de un brote hospitalario de enterobacterias resistentes a carbapenémicos.

Objetivos: Realizar la caracterización microbiológica y molecular de un brote hospitalario de enterobacterias resistentes a carbapenémicos (ERC) en un hospital de tercer nivel en Jalisco, México. Material y Métodos: De septiembre del 2014 a julio del 2015, se colectaron todos los aislados clínicos ERC en el Hospital Civil “Fray Antonio Alcalde” en Guadalajara, Jalisco. Se les determinó la susceptibilidad a antibióticos, la producción de carbapenemasas y se investigaron los genes que codifican para las enzimas KPC, NDM, IMP, VIM y OXA-48 y los perfiles plasmídicos. Se determinó la capacidad de los aislamientos de transferir la resistencia a carbapenémicos mediante ensayos de conjugación y Southern-blot; la relación clonal mediante electroforesis en gel de campos pulsados (EGCP) y se realizó la tipificación de secuencias multi locus (MLST). Se determinaron la producción de biopelícula y la presencia de 14 genes de virulencia en aislamientos de K. pneumoniae resistentes y susceptibles a carbapenémicos. Resultados: Se detectaron 52 aislamientos resistentes a carbapenémicos: Klebsiella pneumoniae (n = 46), Enterobacter cloacae (n = 3), Escherichia coli (n = 1), Providencia rettgeri (n = 1) y Citrobacter freundii (n = 1). El gen detectado en mayor frecuencia fue NDM-1 (92.3%), seguido de VIM (4%), IMP (2%) y KPC (2%). El gen NDM-1 se detectó en plásmidos de 130 a 170 kb en K. pneumoniae (90%), E. cloacae (6%), E. coli (2%) y P. rettgeri (2%); y se detectó en diferentes fragmentos de restricción. La clona predominante (clona A) de K. pneumoniae incluyó 28/48 (60%) aislamientos. Se detectaron K. pneumoniae ST307, ST309, ST392, ST846, ST2399 y ST2400, así como E. cloacae ST182 y E. coli ST10. Los genes fimA y uge fueron detectados con más frecuencia en cepas de K. pneumoniae susceptibles a carbapemémicos (p = <0.001) y la producción de biopelícula se detectó con más frecuencia en aislamientos resistentes a carbapenémicos (p = <0.05). Conclusiones: En el estudio del brote hospitalario por ERC en el “Hospital Civil de Guadalajara” en Jalisco, México se detectaron cuatro especies de enterobacterias portadoras del gen NDM-1. El brote se propagó por transferencia horizontal del plásmido portador del gen NDM-1, y por transmisión de cepas portadoras del gen. La presencia de plásmidos de distinto tamaño como portadores del gen NDM- 1, así como la presencia de este gen en diferentes fragmentos plasmídicos sugieren un rearreglo de los plásmidos portadores. La presencia de los genes VIM, KPC e IMP se consideró como un evento esporádico

Aztreonam plus ceftazidime-avibactam as treatment of NDM-1-producing Klebsiella pneumoniae bacteraemia in a neutropenic patient: Last resort therapy?

Objectives: We report the successful treatment of a bloodstream infection caused by Klebsiella pneumoniae harbouring NDM-1 using aztreonam-ceftazidime-avibactam in a neutropenic patient in whom colistin and meropenem therapy had previously failed. Methods: A clinical isolate was evaluated to determine the presence of NDM, TEM, SHV, CTX, and CMY, and the killing kinetics of aztreonam (ATM; 4 mg/mL), aztreonam-avibactam (ATM-AVI; 4/4 mg/mL), and colistin (2 and 4 mg/mL) were tested. Results: ATM-AVI showed in vitro activity against the Klebsiella pneumoniae harbouring NDM-1, whereas colistin allowed re-growth. Conclusions: This report supports reconsideration of use of colistin for treatment of infections caused by K. pneumoniae harbouring NDM. CZA/ATM use should be kept in mind as a treatment option, perhaps earlier than colistin

Molecular and microbiological report of a hospital outbreak of NDM-1-carrying Enterobacteriaceae in Mexico

Abstract Objectives To characterize the microbiological, molecular and epidemiological data of an outbreak of carbapenem-resistant Enterobacteriaceae (CRE) in a tertiary-care hospital in Mexico. Methods From September 2014 to July 2015, all CRE clinical isolates recovered during an outbreak in the Hospital Civil "Fray Antonio Alcalde" in Jalisco, Mexico were screened for antimicrobial susceptibility, carbapenemase production, carbapenemase-encoding genes, and plasmid profiles. Horizontal transfer of imipenem resistance; and clonal diversity by pulsed-field gel electrophoresis (PFGE) and multilocus sequence typing (MLST); as well as biofilm production and the presence of 14 virulence genes were analyzed in selected isolates. Results Fifty-two carbapenem-resistant isolates corresponding to 5 species were detected, i.e., Klebsiella pneumoniae (n = 46), Enterobacter cloacae (n = 3), Escherichia coli (n = 1), Providencia rettgeri (n = 1) and Citrobacter freundii (n = 1) with carbapenemase encoding genes blaNDM-1 (n = 48), blaVIM (n = 3), blaIMP (n = 1) and blaKPC (n = 1) detected in these isolates. The blaNDM-1 gene was detected in plasmids from 130- to 170-kb in K. pneumoniae (n = 46); E. cloacae (n = 3), E. coli (n = 1) and P. rettgeri (n = 1). The transfer of plasmids harboring the blaNDM-1 gene was obtained in eight transconjugants. One plasmid restriction pattern was detected, with the blaNDM-1 identified in different restriction fragments. Predominant clone A of K. pneumoniae isolates archived 28/46 (60%) isolates and belongs to ST392. Besides, ST307, ST309, ST846, ST2399, and ST2400 were detected for K. pneumoniae; as well as E. cloacae ST182 and E. coli ST10. The fimA and uge genes were more likely to be identified in K. pneumoniae carbapenemsusceptible isolates (p =<0.001) and biofilm production was more liable to be observed in carbapenem-resistant isolates (p =<0.05). Conclusions Four Enterobacteriaceae species harboring the blaNDM-1 gene were detected in a nosocomial outbreak in Mexico; horizontal transfer and strain transmission were demonstrated for the blaNDM-1 gene. Given the variation in the size of the plasmid harboring blaNDM-1, complex rearrangements must also be occurring

Risk factors and outcome associated with the acquisition of linezolid-resistant Enterococcus faecalis

Objectives: Linezolid is a synthetic oxazolidinone antibiotic frequently used to treat vancomycin-resistant enterococcal infections. Vancomycin-susceptible Enterococcus faecalis can develop resistance to linezolid in environments with excessive linezolid use. The aim of this study was to define risk factors and outcome associated with the acquisition of linezolid-resistant E. faecalis (LREfs). Methods: A retrospective case–control study was designed including patients hospitalised from January 2014 to October 2017 at Hospital Civil de Guadalajara ‘Fray Antonio Alcalde’ in Guadalajara, Mexico. A total of 50 patients culture-positive for LREfs and 100 control patients hospitalised in the same room and time as the cases were included. Clinical and demographic data were collected and analysed. Results: Risk factors for the presence of LREfs included prior linezolid use [odds ratio (OR) = 6.74], prior clindamycin use (OR = 6.72) and previous surgery (OR = 5.79). The mortality rate was 18% for LREfs cases versus 9% for controls. Conclusion: LREfs has emerged and spread in our hospital, an environment in which linezolid use is considerable. Risk factors for LREfs are prior antibiotic use, including linezolid, and previous surgery

Risk factors and molecular mechanisms associated with trimethoprim–sulfamethoxazole resistance in Stenotrophomonas maltophilia in Mexico

Abstract Purpose. Stenotrophomonas maltophilia is a multidrug-resistant opportunistic pathogen causing an increasing number of nosocomial infections. Our aim was to evaluate the risk factors and mechanisms associated with trimethoprim– sulfamethoxazole (SXT) resistance in S. maltophilia infections in Mexico. Methodology. Clinical isolates and patients’ demographic and clinical data were collected from February 2007 to August 2015 in two tertiary-care hospitals in Mexico. Antimicrobial susceptibility and analysis of sul and SmeABC and SmeDEF efflux pump overexpression were performed in all isolates. Results/Key findings. In the 9-year period, 196 patients infected with S. maltophilia were identified. Most patients were male, and the mean age was 46.2years. The mean Charlson score was 1.42, and the most frequent comorbidities were arterial hypertension (26.7%), type 2 diabetes (21.2%) and cerebral infarction (11.6%). High drug resistance to meropenem (93.4%), gentamicin (55.1%), ceftazidime (52.3%), cefotaxime (51.5%), amikacin (42.3%) and cefepime (32.1%), and lower resistance to ciprofloxacin (26.0%), SXT (25.0%), chloramphenicol (14.3%) and levofloxacin (2.6%) were detected. SXT resistance was not associated with the sul genes. SmeABC overexpression was associated with gentamicin (P=0.001) and levofloxacin resistance (P=0.041), whereas SmeDEF overexpression was associated with ceftazidime resistance (P=0.003). Prolonged hospitalization (�15days) was an independent risk factor for SXT-resistant S. maltophilia infections (OR=3.05; 95%CI=1.12– 8.86; P=0.029). Conclusion. Given the high SXT resistance rate, SXT is not an effective first-line therapy for our patients; instead, levofloxacin could be used as an appropriate therapeutic option against S. maltophilia infections

The carriage of interleukin-1B-31*C allele plus Staphylococcus aureus and Haemophilus influenzae increases the risk of recurrent tonsillitis in a Mexican population

Abstract The aim of the present study was to estimate the relative contribution of immunogenetic and microbiological factors in the development of recurrent tonsillitis in a Mexican population. Patients (n = 138) with recurrent tonsillitis and an indication of tonsillectomy (mean age: 6.05 years±3.00; median age: 5 years, female: 58; age range: 1–15 years) and 195 nonrelated controls older than 18 years and a medical history free of recurrent tonsillitis were included. To evaluate the microbial contribution, tonsil swab samples from both groups and extracted tonsil samples from cases were cultured. Biofilm production of isolated bacteria was measured. To assess the immunogenetic component, DNA from peripheral blood was genotyped for the TNFA-308G/A single-nucleotide polymorphism (SNP) and for the IL1B -31C/T SNP. Normal microbiota, but no pathogens or potential pathogens, were identified from all control sample cultures. The most frequent pathogenic species detected in tonsils from cases were Staphylococcus aureus (48.6%, 67/138) and Haemophilus influenzae (31.9%, 44/138), which were found more frequently in patient samples than in samples from healthy volunteers (P<0.0001). Importantly, 41/54 (75.9%) S. aureus isolates were biofilm producers (18 weak and 23 strong), whereas 17/25 (68%) H. influenzae isolates were biofilm producers (10 weak, and 7 strong biofilm producers). Patients with at least one copy of the IL1B-31*C allele had a higher risk of recurrent tonsillitis (OR = 4.03; 95% CI = 1.27– 14.27; P = 0.013). TNFA-308 G/A alleles were not preferentially distributed among the groups. When considering the presence of IL1B-31*C plus S. aureus, IL1B-31*C plus S. aureus biofilm producer, IL1B-31*C plus H. influenzae or IL1B-31*C plus H. influenzae biofilm producer, the OR tended to infinite. Thus, the presence of IL1B-31*C allele plus the presence of S. aureus and/or H. influenzae could be related to the development of tonsillitis in this particular Mexican population

The successful containment of a hospital outbreak caused by NDM-1-producing Klebsiella pneumoniae ST307 using active surveillance

The worldwide dissemination of high-risk carbapenemase-producing Klebsiella pneumoniae clones has become a major threat to healthcare facilities. This study describes the successful containment of a hospital outbreak caused by NDM-1-producing K. pneumoniae Sequence Type (ST) 307 using active surveillance. The outbreak began when a patient was transferred from a local hospital. After 48 hours in our hospital, a tracheal aspirate was positive for a meropenem resistant and carbapenemase-producing K. pneumoniae. All patients in the medical intensive care unit (ICU) and the neurology wards were subject to contact precautions. The hospital surfaces and devices, healthcare workers, and patients from these wards were screened by cultures. Fecal swabs were placed into broth and PCR for blaKPC, blaOXA-48, blaIMP, blaVIM, and blaNDM, which were performed directly from the broth after 12 hours. PCRs were also performed on DNA extracted from carbapenemase-producing species from subcultured broths. Five and nine days later, two more patients’ rectal swabs tested positive. Molecular assays identified K. pneumoniae blaNDM-1 onto a 130-kb conjugative plasmid (IncY, IncFIIs, and IncFIIY), ST307. After the three patients were discharged, monitoring continued, and after three weeks with negative results, rectal swabbing ended. In conclusion, it was possible to contain a hospital outbreak caused by NDM-1-producing K. pneumoniae ST307 through epidemiological and microbiological surveillance. With the methodology used, the detection of NDM-type genes in fecal samples was obtained in approximately 15 hours after obtaining the fecal sample

A case–control study of infections caused by Klebsiella pneumoniae producing New Delhi metallo-beta-lactamase-1: Predictors and outcomes

IntroductionInfections caused by antimicrobial-resistant bacteria are a significant cause of death worldwide, and carbapenemase-producing bacteria are the principal agents. New Delhi metallo-beta-lactamase-1 producing Klebsiella pneumoniae (KP-NDM-1) is an extensively drug-resistant bacterium that has been previously reported in Mexico. Our aim was to conduct a case–control study to describe the risk factors associated with nosocomial infections caused by K. pneumoniae producing NDM-1 in a tertiary-care hospital in Mexico.MethodsA retrospective case–control study with patients hospitalized from January 2012 to February 2018 at the Hospital Civil de Guadalajara “Fray Antonio Alcalde” was designed. During this period, 139 patients with a culture that was positive for K. pneumoniae NDM-1 (cases) and 486 patients hospitalized in the same department and on the same date as the cases (controls) were included. Data were analyzed using SPSS v. 24, and logistic regression analysis was conducted to calculate the risk factors for KP-NDM-1 infection.ResultsOne hundred and thirty-nine case patients with a KP-NDM-1 isolate and 486 control patients were analyzed. In the case group, acute renal failure was a significant comorbidity, hospitalization days were extended, and significantly more deaths occurred. In a multivariate analysis of risk factors, the independent variables included the previous use of antibiotics (odds ratio, OR = 12.252), the use of a urinary catheter (OR = 5.985), the use of a central venous catheter (OR = 5.518), the use of mechanical ventilation (OR = 3.459), and the length of intensive care unit (ICU) stay (OR = 2.334) as predictors of infection with NDM-1 K. pneumoniae.ConclusionIn this study, the previous use of antibiotics, the use of a urinary catheter, the use of a central venous catheter, the use of mechanical ventilation, and ICU stay were shown to be predictors of infection with NDM-1 K. pneumoniae and were independent risk factors for infection with NDM-1 K. pneumoniae

Species identification of Enterococcus

Objetivo: Aquí, evaluamos el rendimiento de dos sistemas comerciales de EM MALDI ‐ TOF y tres sistemas basados en bioquímicos y los comparamos con WGS como el estándar de oro para identificar cepas de enterococos resistentes a la vancomicina (ERV). Métodos: Se incluyeron un total de 87 aislamientos clínicos de ERV. Los espectrómetros de masas fueron el sistema Microflex con software Biotyper 3.1 y el sistema Vitek MS. Los sistemas basados en bioquímicos incluyeron los sistemas Vitek 2, Phoenix y MicroScan WalkAway. La WGS se realizó en un instrumento Illumina MiSeq utilizando el kit de reactivos MiSeq v3. La resistencia a la vancomicina se determinó según los criterios de CLSI. Resultados: Entre los 87 VRE, 71 y 16 fueron identificados como Enterococcus faecium y Enterococcus faecalis por WGS. Los 71 E faecium fueron identificados correctamente por ambos espectrómetros de masas, así como por los instrumentos Vitek 2 y Phoenix. Sin embargo, el sistema MicroScan solo identificó correctamente 51 aislados de E. faecium . La identificación errónea más frecuente fue Enterococcus casseliflavus (n = 20). Para E. faecium resistente a la vancomicina , el sistema Microflex Biotyper tuvo la mayor sensibilidad (85,54%) y todos los instrumentos (excepto el Microscan) tuvieron una especificidad y un VPP del 100%. Hasta el 87% de E. faecalis los aislamientos fueron identificados erróneamente por VITEK MS y VITEK2, 81% por Microscan y Phoenix, y 75% por Bruker biotyper. Conclusión: A medida que la cobertura de la base de datos de secuencias de tipo cepa-genoma continúa creciendo y el costo de la secuenciación del ADN continúa disminuyendo, la identificación basada en el genoma puede ser una herramienta útil para los laboratorios de diagnóstico, con su precisión superior incluso sobre MALDI-TOF y las operaciones basadas en bases de datos .Aim: Here, we evaluated the performance of two commercial MALDI-TOF MS systems and three biochemical-based systems and compared them to WGS as the gold standard for identifying isolates of vancomycin-resistant enterococci (VRE). Methods: A total of 87 VRE clinical isolates were included. The mass spectrometers were the Microflex system with Biotyper software 3.1 and the Vitek MS system. The biochemical-based systems included the Vitek 2, Phoenix, and MicroScan WalkAway systems. WGS was performed on an Illumina MiSeq instrument using the MiSeq v3 reagent kit. Vancomycin resistance was determined according to CLSI criteria. Results: Among the 87 VRE, 71 and 16 were identified as Enterococcus faecium and Enterococcus faecalis by WGS. All 71 E faecium were correctly identified by both mass spectrometers, as well as the Vitek 2 and Phoenix instruments. However, only 51 E faecium isolates were correctly identified by the MicroScan system. The most frequent misidentification was Enterococcus casseliflavus (n = 20). For vancomycin-resistant E faecium, the Microflex Biotyper system had the highest sensitivity (85.54%), and all instruments (except for the Microscan) had a 100% specificity and PPV. Up to 87% of E faecalis isolates were misidentified by VITEK MS and VITEK2, 81% by Microscan and Phoenix, and 75% by Bruker biotyper. Conclusion: As the coverage of type strain-genome sequence database continues to grow and the cost of DNA sequencing continues to decrease, genome-based identification can be a useful tool for diagnostic laboratories, with its superior accuracy even over MALDI-TOF and database-driven operations