DSpace Repository :: Browsing by Author "Diaz, Lorena"

Browsing by Author "Diaz, Lorena"

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A liaF codon deletion abolishes daptomycin bactericidal activity against vancomycin-resistant enterococcus faecali
(American Society for Microbiology, 2013) Munita, José; Tran, Truc; Diaz, Lorena; Panesso, Diana; Reyes, Jinnethe; Murray, Barbara; Arias, Cesar
The genetic bases for antibiotic tolerance are obscure. Daptomycin (DAP) is a lipopeptide antibiotic with bactericidal activity against enterococci. Using time-kill assays, we provide evidence for the first time that a deletion of isoleucine in position 177 of LiaF, a member of the three-component regulatory system LiaFSR involved in the cell envelope response to antimicrobials, is directly responsible for a DAP-tolerant phenotype and is likely to negatively affect response to DAP therapy.
A prospective cohort multicenter study of molecular epidemiology and phylogenomics of Staphylococcus aureus bacteremia in nine Latin American countries
(2017) Arias, Cesar A; Reyes, Jinnethe; Carvajal, Lina; Rincon, Sandra; Diaz, Lorena; Panesso, Diana; Ibarra, Gabriel; Rios, Rafael; Munita, José; Salles, Mauro J; Alvarez-Moreno, Carlos; Labarca, Jaime; Garcia, Coralith; Luna, Carlos M; Mejia-Villatoro, Carlos; Zurita, Jeanette; Guzman-Blanco, Manuel; Rodriguez-Noriega, Eduardo; Narechania, Apurva; Rojas, Laura J; Planet, Paul J; Weinstock, George M; Gotuzzo, Eduardo; Seasg, Carlos
Staphylococcus aureus is an important pathogen causing a spectrum of diseases ranging from mild skin and soft tissue infections to life-threatening conditions. Bloodstream infections are particularly important, and the treatment approach is complicated by the presence of methicillin-resistant S. aureus (MRSA) isolates. The emergence of new genetic lineages of MRSA has occurred in Latin America (LA) with the rise and dissemination of the community-associated USA300 Latin American variant (USA300-LV). Here, we prospectively characterized bloodstream MRSA recovered from selected hospitals in 9 Latin American countries. All isolates were typed by pulsed-field gel electrophoresis (PFGE) and subjected to antibiotic susceptibility testing. Whole-genome sequencing was performed on 96 MRSA representatives. MRSA represented 45% of all (1,185 S. aureus) isolates. The majority of MRSA isolates belonged to clonal cluster (CC) 5. In Colombia and Ecuador, most isolates (72%) belonged to the USA300-LV lineage (CC8). Phylogenetic reconstructions indicated that MRSA isolates from participating hospitals belonged to three major clades. Clade A grouped isolates with sequence type 5 (ST5), ST105, and ST1011 (mostly staphylococcal chromosomal cassette mec [SCCmec] I and II). Clade B included ST8, ST88, ST97, and ST72 strains (SCCmec IV, subtypes a, b, and c/E), and clade C grouped mostly Argentinian MRSA belonging to ST30. In summary, CC5 MRSA was prevalent in bloodstream infections in LA with the exception of Colombia and Ecuador, where USA300-LV is now the dominant lineage. Clonal replacement appears to be a common phenomenon, and continuous surveillance is crucial to identify changes in the molecular epidemiology of MRSA.
Ceftaroline-resistant, daptomycin-tolerant, and heterogeneous vancomycin-intermediate methicillin-resistant staphylococcus aureus causing infective endocarditis
(American Society for Microbiology, 2017) Nigo, Masayuki; Diaz, Lorena; Carvajal, Lina P.; Tran, Truc T.; Rios, Rafael; Panesso, Diana; Garavito, Juan D; Miller, William; Wagner, Audrey
We report a case of infective endocarditis (IE) caused by ceftaroline-resistant, daptomycin-tolerant, and heterogeneous vancomycin-intermediate methicillin-resistant S. aureus (MRSA). Resistance to ceftaroline emerged in the absence of drug exposure, and the E447K substitution in the active site of PBP2a previously associated with ceftaroline resistance was identified. Additionally, we present evidence of patient-to-patient transmission of the strain within the same unit. This case illustrates the difficulties in treating MRSA IE in the setting of a multidrug-resistant phenotype.
Clinical characteristics, microbiology and outcomes of a cohort of patients treated with ceftolozane/tazobactam in acute care inpatient facilities, Houston, Texas, USA
(2023) Tran, Truc; Cabrera, Nicolo; Gonzales, Anne; Carlson, Travis; Alnezary, Faris; Miller, William; Sakurai, Aki; Dinh, An; Rydell, Kirsten; Rios, Rafael; Diaz, Lorena; Hanson, Blake; Munita, Jose M.; Pedroza, Claudia; Shelburne, Samuel; Aitken, Samuel; Garey, Kevin; Dillon, Ryan; Puzniak, Laura; Arias, Cesar
Background: Ceftolozane/tazobactam is a β-lactam/β-lactamase inhibitor combination with activity against a variety of Gram-negative bacteria, including MDR Pseudomonas aeruginosa. This agent is approved for hospital-acquired and ventilator-associated bacterial pneumonia. However, most real-world outcome data come from small observational cohorts. Thus, we sought to evaluate the utilization of ceftolozane/tazobactam at multiple tertiary hospitals in Houston, TX, USA. Methods: We conducted a multicentre retrospective study of patients receiving at least 48 h of ceftolozane/tazobactam therapy from January 2016 through to September 2019 at two hospital systems in Houston. Demographic, clinical and microbiological data were collected, including the infecting bacterial isolate, when available. The primary outcome was composite clinical success at hospital discharge. Secondary outcomes included in-hospital mortality and clinical disposition at 14 and 30 days post ceftolozane/tazobactam initiation. Multivariable logistic regression analysis was used to identify predictors of the primary outcome and mortality. Recovered isolates were tested for susceptibility to ceftolozane/tazobactam and underwent WGS. Results: A total of 263 patients were enrolled, and composite clinical success was achieved in 185 patients (70.3%). Severity of illness was the most consistent predictor of clinical success. Combination therapy with ceftolozane/tazobactam and another Gram-negative-active agent was associated with reduced odds of clinical success (OR 0.32, 95% CI 0.16-0.63). Resistance to ceftolozane/tazobactam was noted in 15.4% of isolates available for WGS; mutations in ampC and ftsI were common but did not cluster with a particular ST. Conclusions: Clinical success rate among this patient cohort treated with ceftolozane/tazobactam was similar compared with previous experiences. Ceftolozane/tazobactam remains an alternative agent for treatment of susceptible isolates of P. aeruginosa.
Correlation between mutations in liaFSR of enterococcus faecium and MIC of daptomycin: revisiting daptomycin breakpoints
(American Society for Microbiology, 2012) Munita, José; Panesso, Diana; Diaz, Lorena; Tran, Truc; Reyes, Jinnethe; Wanger, Audrey; Murray, Barbara; Arias, Cesar
Mutations in liaFSR, a three-component regulatory system controlling cell-envelope stress response, were recently linked with the emergence of daptomycin (DAP) resistance in enterococci. Our previous work showed that a liaF mutation increased the DAP MIC of a vancomycin-resistant Enterococcus faecalis strain from 1 to 3 μg/ml (the DAP breakpoint is 4 μg/ml), suggesting that mutations in the liaFSR system could be a pivotal initial event in the development of DAP resistance. With the hypothesis that clinical enterococcal isolates with DAP MICs between 3 and 4 μg/ml might harbor mutations in liaFSR, we studied 38 Enterococcus faecium bloodstream isolates, of which 8 had DAP MICs between 3 and 4 μg/ml by Etest in Mueller-Hinton agar. Interestingly, 6 of these 8 isolates had predicted amino acid changes in the LiaFSR system. Moreover, we previously showed that among 6 DAP-resistant E. faecium isolates (MICs of >4 μg/ml), 5 had mutations in liaFSR. In contrast, none of 16 E. faecium isolates with a DAP MIC of ≤2 μg/ml harbored mutations in this system (P < 0.0001). All but one isolate with liaFSR changes exhibited DAP MICs of ≥16 μg/ml by Etest using brain heart infusion agar (BHIA), a medium that better supports enterococcal growth. Our findings provide a strong association between DAP MICs within the upper susceptibility range and mutations in the liaFSR system. Concomitant susceptibility testing on BHIA may be useful for identifying these E. faecium first-step mutants. Our results also suggest that the current DAP breakpoint for E. faecium may need to be reevaluated.
Daptomycin-resistant enterococcus faecalis diverts the antibiotic molecule from the division septum and remodels cell membrane phospholipids
(American Society for Microbiology, 2013) Tran, Truc; Panesso, Diana; Mishra, Nagendra; Mileykovskaya, Eugenia; Guan, Ziqianq; Munita, José; Reyes, Jinnethe; Diaz, Lorena; Weinstock, George; Murray, Barbara; Shamoo, Yousif; Dowhan, William; Bayer, Arnold; Arias, Cesar
Treatment of multidrug-resistant enterococci has become a challenging clinical problem in hospitals around the world due to the lack of reliable therapeutic options. Daptomycin (DAP), a cell membrane-targeting cationic antimicrobial lipopeptide, is the only antibiotic with in vitro bactericidal activity against vancomycin-resistant enterococci (VRE). However, the clinical use of DAP against VRE is threatened by emergence of resistance during therapy, but the mechanisms leading to DAP resistance are not fully understood. The mechanism of action of DAP involves interactions with the cell membrane in a calcium-dependent manner, mainly at the level of the bacterial septum. Previously, we demonstrated that development of DAP resistance in vancomycin-resistant Enterococcus faecalis is associated with mutations in genes encoding proteins with two main functions, (i) control of the cell envelope stress response to antibiotics and antimicrobial peptides (LiaFSR system) and (ii) cell membrane phospholipid metabolism (glycerophosphoryl diester phosphodiesterase and cardiolipin synthase). In this work, we show that these VRE can resist DAP-elicited cell membrane damage by diverting the antibiotic away from its principal target (division septum) to other distinct cell membrane regions. DAP septal diversion by DAP-resistant E. faecalis is mediated by initial redistribution of cell membrane cardiolipin-rich microdomains associated with a single amino acid deletion within the transmembrane protein LiaF (a member of a three-component regulatory system [LiaFSR] involved in cell envelope homeostasis). Full expression of DAP resistance requires additional mutations in enzymes (glycerophosphoryl diester phosphodiesterase and cardiolipin synthase) that alter cell membrane phospholipid content. Our findings describe a novel mechanism of bacterial resistance to cationic antimicrobial peptides. IMPORTANCE: The emergence of antibiotic resistance in bacterial pathogens is a threat to public health. Understanding the mechanisms of resistance is of crucial importance to develop new strategies to combat multidrug-resistant microorganisms. Vancomycin-resistant enterococci (VRE) are one of the most recalcitrant hospital-associated pathogens against which new therapies are urgently needed. Daptomycin (DAP) is a calcium-decorated antimicrobial lipopeptide whose target is the bacterial cell membrane. A current paradigm suggests that Gram-positive bacteria become resistant to cationic antimicrobial peptides via an electrostatic repulsion of the antibiotic molecule from a more positively charged cell surface. In this work, we provide evidence that VRE use a novel strategy to avoid DAP-elicited killing. Instead of "repelling" the antibiotic from the cell surface, VRE diverts the antibiotic molecule from the septum and "traps" it in distinct membrane regions. We provide genetic and biochemical bases responsible for the mechanism of resistance and disclose new targets for potential antimicrobial development.
Dissecting the mechanisms of linezolid resistance in a drosophila melanogaster infection model of staphylococcus aureus
(Oxford University Press, 2013) Diaz, Lorena; Kontoyiannis, Dimitrios; Panesso, Diana; Albert, Nathaniel; Singh, Kavindra; Tran, Truc; Munita, José; Murray, Barbara; Arias, Cesar
BACKGROUND: Mini-host models are simple experimental systems to study host-pathogen interactions. We adapted a Drosophila melanogaster infection model to evaluate the in vivo effect of different mechanisms of linezolid (LNZ) resistance in Staphylococcus aureus. METHODS: Fly survival was evaluated after infection with LNZ-resistant S. aureus strains NRS119 (which has mutations in 23S ribosomal RNA [rRNA]), CM-05 and 004-737X (which carry cfr), LNZ-susceptible derivatives of CM-05 and 004-737X (which lack cfr), and ATCC 29213 (an LNZ-susceptible control). Flies were then fed food mixed with LNZ (concentration, 15-500 µg/mL). Results were compared to those in mouse peritonitis, using LNZ via oral gavage at 80 and 120 mg/kg every 12 hours. RESULTS: LNZ at 500 µg/mL in fly food protected against all strains, while concentrations of 15-250 µg/mL failed to protect against NRS119 (survival, 1.6%-20%). An in vivo effect of cfr was only detected at concentrations of 30 and 15 µg/mL. In the mouse peritonitis model, LNZ (at doses that mimic human pharmacokinetics) protected mice from challenge with the cfr+ 004-737X strain but was ineffective against the NRS119 strain, which carried 23S rRNA mutations. CONCLUSIONS: The fly model offers promising advantages to dissect the in vivo effect of LNZ resistance in S. aureus, and findings from this model appear to be concordant with those from the mouse peritonitis model.
Failure of High-Dose Daptomycin for Bacteremia Caused by Daptomycin-Susceptible Enterococcus faecium Harboring LiaSR Substitutions
(2014) Munita, José; Mishra, Nagendra N.; Alvarez, Danya; Tran, Truc T; Diaz, Lorena; Panesso, Diana; Reyes, Jinnethe; Murray, Barbara E; Adachi, Javier A; Bayer, Arnold S; Arias, Cesar A
High-dose daptomycin (DAP) therapy failed in a neutropenic patient with bloodstream infection caused by a DAP-susceptible Enterococcus faecium (minimum inhibitory concentration, 3 µg/mL) harboring genetic changes associated with DAP resistance, with persistent bacteremia and selection of additional resistances. Daptomycin monotherapy should be used cautiously against DAP-susceptible E. faecium strains with minimum inhibitory concentrations >2 µg/mL.
Geographic divergence of methicillin-resistant Staphylococcus aureus ST5-SCCmecI in the aftermath of a major earthquake and tsunami: impact of a plasmid harboring heavy metal resistance genes
(2025) Martínez, Jose; Alcalde, Manuel; Jara, Estefanía; Reyes, Jinnethe; Carvajal, Lina; Rincon, Sandra; Ríos, Rafael; Diaz, Lorena; Quesille, Ana; Riquelme, Roberto; Rivas Jiménez, Lina María; Moustafa, Ahmed; Hanson, Blake; Undurraga, Eduardo; Olivares, Jorge; García, Patricia; Araos Bralic, Rafael Ignacio; Planet, Paul; Arias, César; Munita, Jose M.
El Staphylococcus aureus resistente a la meticilina (SARM) es una importante amenaza para la salud pública. La propagación global del SARM se caracteriza por sucesivas oleadas de clones epidémicos que dominan regiones geográficas específicas. Se cree que la adquisición de genes que codifican la resistencia a metales pesados (GMMP) es una característica clave en la divergencia geográfica del SARM. Sin embargo, la relación causa-efecto entre la presencia de GMMP y la divergencia de los clones de SARM aún no se ha dilucidado. En este estudio, evaluamos el papel que los GMMP pudieron haber desempeñado en la divergencia evolutiva del linaje ST5-SCC mec I del SARM en Latinoamérica. Realizamos una caracterización genómica de 113 aislamientos clínicos de SARM de seis centros de salud latinoamericanos, incluyendo 53 aislamientos recolectados en dos ciudades de Chile (Santiago y Concepción). Encontramos un plásmido (pSCL4752) que alberga genes de resistencia al arsénico, cadmio y mercurio en el 65% ( n = 71) de los aislados ST5-SCC mec I. También observamos una divergencia geográfica asociada a la presencia de pSCL4752 en aislados chilenos, con una mayor frecuencia en aislados de Concepción (88%) en comparación con Santiago (29%). Curiosamente, un análisis de reloj molecular reveló que esta divergencia se produjo tras el terremoto y tsunami de magnitud 8,8 Mw que azotó la zona de Concepción en 2010. Además, nuestros resultados demuestran que la presencia de pSCL4752 puede ser beneficiosa o perjudicial para los aislados ST5-SCC mec I, dependiendo de la disponibilidad ambiental de estos metales pesados. Nuestros resultados sugieren que la divergencia del linaje ST5-SCC mec I MRSA en América Latina podría haber sido fomentada por desastres ambientales e influenciada por la presencia/ausencia de HMRG albergados en un plásmido. IMPORTANCIA El Staphylococcus aureus resistente a la meticilina (MRSA) es una de las principales causas de infecciones potencialmente mortales en todo el mundo y una creciente preocupación para la salud pública. El aumento de bacterias resistentes a los antibióticos, como el MRSA, a menudo está vinculado a adaptaciones genéticas que mejoran su supervivencia. Nuestra investigación arroja luz sobre cómo los cambios ambientales, como los desencadenados por un desastre natural, pueden influir en la evolución y la propagación geográfica de un linaje de MRSA altamente resistente en América Latina. Identificamos un plásmido que porta genes de resistencia al arsénico, cadmio y mercurio, el cual se asoció con la divergencia geográfica del ST5-SCC mec.Se ha identificado un linaje de SARM con diferencias notables en su prevalencia entre las regiones afectadas por un gran terremoto y tsunami. Al vincular los eventos ambientales con la evolución del patógeno, nuestro estudio destaca el papel de las presiones ecológicas en la propagación del SARM. Estos hallazgos subrayan la necesidad de integrar el monitoreo ambiental en las estrategias de salud pública para comprender mejor el desafío global de la resistencia a los antimicrobianos.
Membrane Lipids Augment Cell Envelope Stress Signaling via the MadRS System to Defend Against Antimicrobial Peptides and Antibiotics in Enterococcus faecalis
(2025) Miller, William; Nguyen, April; Singh, Kavindra; Rizvi, Samie; Khan, Ayesha; Erickson, Sam; Egge, Stephanie; Cruz, Melissa; Dinh, An; Diaz, Lorena; Thornton, Philip; Zhang, Rutan; Xu, Libin; Garsin, Danielle; Shamoo, Yousif; Arias, Cesar
Enterococci have evolved resistance mechanisms to protect their cell envelopes against bacteriocins and host cationic antimicrobial peptides (CAMPs) produced in the gastrointestinal environment. Activation of the membrane stress response has also been tied to resistance to the lipopeptide antibiotic daptomycin. However, the actual effectors mediating resistance have not been elucidated. Here, we show that the MadRS (formerly YxdJK) membrane antimicrobial peptide defense system controls a network of genes, including a previously uncharacterized 3-gene operon (madEFG) that protects the Enterococcus faecalis cell envelope from antimicrobial peptides. Constitutive activation of the system confers protection against CAMPs and daptomycin in the absence of a functional LiaFSR system and leads to persistence of cardiac microlesions in vivo. Moreover, changes in the lipid cell membrane environment alter CAMP susceptibility and expression of the MadRS system. Thus, we provide a framework supporting a multilayered envelope defense mechanism for resistance and survival coupled to virulence.
Methicillin-Susceptible, Vancomycin-Resistant Staphylococcus aureus, Brazil
(Centers for Disease Control and Prevention, 2015) Panesso, Diana; Planet, Paul; Diaz, Lorena; Hugonnet, Jean-Emmanuel; Tran, Truc; Narechania, Apurva; Munita, José; Rincon, Sandra; Carvajal, Lina; Reyes, Jinnethe; Londoño, Alejandra; Smith, Hannah; Sebra, Robert; Deikus, Gintaras; Weinstock, George; Murray, Barbara; Rossi, Flavia; Arthur, Michel; Arias, Cesar
We report characterization of a methicillin-susceptible, vancomycin-resistant bloodstream isolate of Staphylococcus aureus recovered from a patient in Brazil. Emergence of vancomycin resistance in methicillin-susceptible S. aureus would indicate that this resistance trait might be poised to disseminate more rapidly among S. aureus and represents a major public health threat.
Multispecies emergence of dual blaKPC/NDM carbapenemase-producing Enterobacterales recovered from invasive infections in Chile
(2025) Quesille, Ana; Solar, Camila; Martínez, Jose; Rivas Jiménez, Lina María; Quiroz, Valeria; González, Ana; Riquelme, Roberto; Ugalde, Juan; Peters, Anne; Ortega, Oscar; Araos Bralic, Rafael Ignacio; García, Patricia; Lebreton, Francois; Munita, Jose M.; Diaz, Lorena
Carbapenemase-producing carbapenem-resistant Enterobacterales (CP-CRE) represent a significant global threat. The emergence of dual CP-CRE is particularly alarming, as they can potentially compromise the efficacy of newer antibiotics, further decreasing therapeutic alternatives. Herein, we report the emergence of multiple species of CP-CRE recovered from invasive infections in Chile that simultaneously harbor blaKPC and blaNDM and provide an in-depth genomic characterization of these worrisome pathogens. We collected carbapenem-resistant Enterobacterales (CRE) isolates from invasive infections over a 4-year period, across 11 healthcare centers in Chile. Bacterial species and the presence of carbapenemase genes were confirmed using MALDI-TOF and PCR assays, respectively. Antimicrobial susceptibility testing was conducted through disk diffusion and broth microdilution methods. Dual CP-CRE isolates were subjected to short- and long-read whole genome sequencing to perform a detailed genomic characterization of the isolates and of the mobile genetic elements harboring the enzymes. From a total of 1,335 CRE isolates, we observed an increase in the prevalence of CP-CRE, from 11% in 2019 to 38% in 2022. A total of 11 dual CP-CRE isolates were recovered, all of them harboring blaKPC and blaNDM. Species corresponded to Escherichia coli (n = 6), Klebsiella pneumoniae (n = 2), Klebsiella oxytoca (n = 2), and Citrobacter freundii (n = 1). Dual CP-CRE isolates exhibited resistance to all tested β-lactams except for cefiderocol. The blaKPC and blaNDM encoding genes were located on independent plasmids. Platforms harboring blaKPC were diverse and included IncN, IncF, and IncFIB plasmids. In contrast, blaNDM-7 was only found on fairly conserved IncX3 plasmids. We report that a rapid increase of CP-CRE in Chile, alongside with the emergence of multiple bacterial species of CP-CRE co-harboring blaKPC-2/3 and blaNDM-7, underscores a critical public health challenge. Our data suggest that the dissemination of blaNDM-7 was predominantly facilitated by IncX3 plasmids, whereas the spread of blaKPC involved multiple plasmid backbones. Active surveillance and genomic monitoring are critical to inform public policy and curtail the spread of these highly resistant pathogens.
Resistance to last resort antibiotics in gram-positive cocci: the post-vancomycin era
(Instituto Nacional de Salud de Colombia, 2014) Rincon, Sandra; Panesso, Diana; Diaz, Lorena; Carvajal, Lina; Reyes, Jinnethe; Munita, José; Arias, Cesar
New therapeutic alternatives have been developed in the last years for the treatment of multidrug-resistant Gram-positive infections. Infections caused by methicillin-resistant Staphylococcus aureus (MRSA) and vancomycin-resistant enterococci (VRE) are considered a therapeutic challenge due to failures and lack of reliable antimicrobial options. Despite concerns related to the use of vancomycin in the treatment of severe MRSA infections in specific clinical scenarios, there is a paucity of solid clinical evidence that support the use of alternative agents (when compared to vancomycin). Linezolid, daptomycin and tigecycline are antibiotics approved in the last decade and newer cephalosporins (such as ceftaroline and ceftobiprole) and novel glycopeptides (dalvavancin, telavancin and oritavancin) have reached clinical approval or are in the late stages of clinical development. This review focuses on discussing these newer antibiotics used in the "post-vancomycin" era with emphasis on relevant chemical characteristics, spectrum of antimicrobial activity, mechanisms of action and resistance, as well as their clinical utility.
The Cefazolin Inoculum Effect Is Associated With Increased Mortality in Methicillin-Susceptible Staphylococcus aureus Bacteremia
(2018) Miller, William; Seas, Carlos; Carvajal, Lina; Diaz, Lorena; Echeverri, Aura; Ferro, Carolina; Rios, Rafael; Porras, Paola; Luna, Carlos; Gotuzzo, Eduardo; Munita, José; Nannini, Esteban; Carcamo, Cesar; Reyes, Jinnethe; Arias, Cesar
Background: Recent studies have favored the use of cefazolin over nafcillin for the treatment of methicillin-susceptible Staphylococcus aureus (MSSA) bacteremia. The clinical influence of the cefazolin inoculum effect (CzIE) in the effectiveness of cephalosporins for severe MSSA infections has not been evaluated. Methods: We prospectively included patients from 3 Argentinian hospitals with S. aureus bacteremia. Cefazolin minimum inhibitory concentrations (MICs) were determined at standard (105 colony-forming units [CFU]/mL) and high (107 CFU/mL) inoculum. The CzIE was defined as an increase of MIC to ≥16 µg/mL when tested at high inoculum. Whole-genome sequencing was performed in all isolates. Results: A total of 77 patients, contributing 89 MSSA isolates, were included in the study; 42 patients (54.5%) had isolates with the CzIE. In univariate analysis, patients with MSSA exhibiting the CzIE had increased 30-day mortality (P = .034) and were more likely to have catheter-associated or unknown source of bacteremia (P = .033) compared with patients infected with MSSA isolates without the CzIE. No statistically significant difference between the groups was observed in age, clinical illness severity, place of acquisition (community vs hospital), or presence of endocarditis. The CzIE remained associated with increased 30-day mortality in multivariate analysis (risk ratio, 2.65; 95% confidence interval, 1.10-6.42; P = .03). MSSA genomes displayed a high degree of heterogeneity, and the CzIE was not associated with a specific lineage. Conclusions: In patients with MSSA bacteremia where cephalosporins are used as firstline therapy, the CzIE was associated with increased 30-day mortality. Clinicians should be cautious when using cefazolin as firstline therapy for these infections.
Transferable vancomycin resistance in a community-associated MRSA lineage
(Massachusetts Medical Society, 2014) Rossi, Flávia; Diaz, Lorena; Wollam, Aye; Panesso, Diana; Zhou, Yanjiao; Rincon, Sandra; Narechania, Apurva; Xing, Galen; Di Gioia, Thais; Doi, André; Tran, Truc; Reyes, Jinnethe; Munita, José; Carvajal, Lina; Hernandez-Roldan, Alejandra; Brandão, Denise; van der Heijden, Inneke Marie; Murray, Barbara; Planet, Paul; Weinstock, George; Arias, Cesar
We report the case of a patient from Brazil with a bloodstream infection caused by a strain of methicillin-resistant Staphylococcus aureus (MRSA) that was susceptible to vancomycin (designated BR-VSSA) but that acquired the vanA gene cluster during antibiotic therapy and became resistant to vancomycin (designated BR-VRSA). Both strains belong to the sequence type (ST) 8 community-associated genetic lineage that carries the staphylococcal chromosomal cassette mec (SCCmec) type IVa and the S. aureus protein A gene (spa) type t292 and are phylogenetically related to MRSA lineage USA300. A conjugative plasmid of 55,706 bp (pBRZ01) carrying the vanA cluster was identified and readily transferred to other staphylococci. The pBRZ01 plasmid harbors DNA sequences that are typical of the plasmid-associated replication genes rep24 or rep21 described in community-associated MRSA strains from Australia (pWBG745). The presence and dissemination of community-associated MRSA containing vanA could become a serious public health concern.
Whole-Genome Analyses of Enterococcus faecium Isolates with Diverse Daptomycin MICs
(American Society for Microbiology, 2014) Diaz, Lorena; Tran, Truc; Munita, José; Miller, William; Rincon, Sandra; Carvajal, Lina; Wollam, Aye; Reyes, Jinnethe; Panesso, Diana; Rojas, Natalia; Shamoo, Yousif; Murray, Barbara; Weinstock, George; Arias, Cesar
Daptomycin (DAP) is a lipopeptide antibiotic frequently used as a "last-resort" antibiotic against vancomycin-resistant Enterococcus faecium (VRE). However, an important limitation for DAP therapy against VRE is the emergence of resistance during therapy. Mutations in regulatory systems involved in cell envelope homeostasis are postulated to be important mediators of DAP resistance in E. faecium. Thus, in order to gain insights into the genetic bases of DAP resistance in E. faecium, we investigated the presence of changes in 43 predicted proteins previously associated with DAP resistance in enterococci and staphylococci using the genomes of 19 E. faecium with different DAP MICs (range, 3 to 48 μg/ml). Bodipy-DAP (BDP-DAP) binding to the cell membrane assays and time-kill curves (DAP alone and with ampicillin) were performed. Genetic changes involving two major pathways were identified: (i) LiaFSR, a regulatory system associated with the cell envelope stress response, and (ii) YycFGHIJ, a system involved in the regulation of cell wall homeostasis. Thr120 → Ala and Trp73 → Cys substitutions in LiaS and LiaR, respectively, were the most common changes identified. DAP bactericidal activity was abolished in the presence of liaFSR or yycFGHIJ mutations regardless of the DAP MIC and was restored in the presence of ampicillin, but only in representatives of the LiaFSR pathway. Reduced binding of BDP-DAP to the cell surface was the predominant finding correlating with resistance in isolates with DAP MICs above the susceptibility breakpoint. Our findings suggest that genotypic information may be crucial to predict response to DAP plus β-lactam combinations and continue to question the DAP breakpoint of 4 μg/ml.
Whole-genome analysis of a daptomycin-susceptible enterococcus faecium strain and its daptomycin-resistant variant arising during therapy
(American Society for Microbiology, 2013) Tran, Truc; Panesso, Diana; Gao, Hongyu; Roh, Jung; Munita, José; Reyes, Jinnethe; Diaz, Lorena; Lobos, Elizabeth; Shamoo, Yousif; Mishra, Nagendra; Bayer, Arnold; Murray, Barbara; Weinstock, George; Arias, Cesar
Development of daptomycin (DAP) resistance in Enterococcus faecalis has recently been associated with mutations in genes encoding proteins with two main functions: (i) control of the cell envelope stress response to antibiotics and antimicrobial peptides (LiaFSR system) and (ii) cell membrane phospholipid metabolism (glycerophosphoryl diester phosphodiesterase and cardiolipin synthase [cls]). However, the genetic bases for DAP resistance in Enterococcus faecium are unclear. We performed whole-genome comparative analysis of a clinical strain pair, DAP-susceptible E. faecium S447 and its DAP-resistant derivative R446, which was recovered from a single patient during DAP therapy. By comparative whole-genome sequencing, DAP resistance in R446 was associated with changes in 8 genes. Two of these genes encoded proteins involved in phospholipid metabolism: (i) an R218Q substitution in Cls and (ii) an A292G reversion in a putative cyclopropane fatty acid synthase enzyme. The DAP-resistant derivative R446 also exhibited an S333L substitution in the putative histidine kinase YycG, a member of the YycFG system, which, similar to LiaFSR, has been involved in cell envelope homeostasis and DAP resistance in other Gram-positive cocci. Additional changes identified in E. faecium R446 (DAP resistant) included two putative proteins involved in transport (one for carbohydrate and one for sulfate) and three enzymes predicted to play a role in general metabolism. Exchange of the "susceptible" cls allele from S447 for the "resistant" one belonging to R446 did not affect DAP susceptibility. Our results suggest that, apart from the LiaFSR system, the essential YycFG system is likely to be an important mediator of DAP resistance in some E. faecium strains.

Browsing by Author "Diaz, Lorena"

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