In Vitro and In Vivo Activity of Citral in Combination with Amphotericin B, Anidulafungin and Fluconazole against Candida auris Isolates.

Candida auris is an emerging fungal pathogen responsible for hospital outbreaks of invasive candidiasis associated with high mortality. The treatment of these mycoses is a clinical challenge due to the high resistance levels of this species to current antifungal drugs, and alternative therapeutic strategies are needed. In this study, we evaluated the in vitro and in vivo activities of combinations of citral with anidulafungin, amphotericin B or fluconazole against 19 C. auris isolates. The antifungal effect of citral was in most cases similar to the effect of the antifungal drugs in monotherapy. The best combination results were obtained with anidulafungin, with synergistic and additive interactions against 7 and 11 of the 19 isolates, respectively. The combination of 0.06 µg/mL anidulafungin and 64 µg/mL citral showed the best results, with a survival rate of 63.2% in Caenorhabditis elegans infected with C. auris UPV 17-279. The combination of fluconazole with citral reduced the MIC of fluconazole from > 64 to 1-4 µg/mL against 12 isolates, and a combination of 2 µg/mL fluconazole and 64 µg/mL citral was also effective in reducing mortality in C. elegans. Amphotericin B combined with citral, although effective in vitro, did not improve the activity of each compound in vivo.

Assessing pH-dependent activities of virulence factors secreted by Candida albicans.

Candida albicans is an opportunistic pathogen that can thrive under adverse conditions including suboptimal pH, nutrient scarcity, and low levels of oxygen. Its pathogenicity is associated with the production of virulence factors such as extracellular hydrolytic enzymes and toxins. This study was aimed at determining the effect of external pH, substrate nature, and strain origin on protease, lipase, and hemolysin production. To achieve this objective, agar plate assays were performed at pH 5.0, 6.5, and 7.5 with substrates suitable for the detection of each family of enzymes. Moreover, the study was conducted with 20 clinical C. albicans isolates from blood, oral cavity, skin, urine, and vagina. The hydrolytic zones formed around the colonies were further measured to calculate the Ez (enzymatic zone) indexes. We found that detection of proteases in skim milk agar plates was possible for most isolates only at pH 5 (80%) and pH 6.5 (75%), whereas BSA plates could confer protease detection exclusively at pH 5 (80%). Similarly, the percentage of isolates possessing lipolytic activities was higher at pH 5 (90%) than at pH 6.5 (70%) and pH 7.5 (35%). In contrast, hemolytic activities were detected in all isolates at pH 6.5 and 7.5 but not at pH 5. Further analysis revealed that some differences in the detected activities could potentially be attributed to the anatomical origin of these isolates. Collectively, these findings suggest that the pH of the site of infection might be critical for mimicking the microenvironment employed to experimentally discover the key virulence factors.

Abemaciclib in combination with pembrolizumab for HR+, HER2- metastatic breast cancer: Phase 1b study.

This nonrandomized, open-label, multi-cohort Phase 1b study (NCT02779751) investigated the safety and efficacy of abemaciclib plus pembrolizumab with/without anastrozole in patients with hormone receptor-positive (HR+), human epidermal growth factor receptor 2-negative (HER2-) metastatic breast cancer (MBC) without prior CDK4 and 6 inhibitor exposure. Patients were divided into two cohorts: treatment naïve (cohort 1) and pretreated (cohort 2). Patients received abemaciclib plus pembrolizumab with (cohort 1) or without (cohort 2) anastrozole over 21-day cycles. The primary objective was safety, and secondary objectives included efficacy and pharmacokinetics (PK). Cohort 1/2 enrolled 26/28 patients, respectively. Neutropenia (30.8/28.6%), AST increase (34.6/17.9%), ALT increase (42.3/10.7%), and diarrhea (3.8/10.7%) were the most frequent grade ≥3 adverse events in cohort 1/2, respectively. A total of two deaths occurred, which investigators attributed to treatment-related adverse events (AEs), both in cohort 1. Higher rates of all grade and grade ≥3 interstitial lung disease (ILD)/pneumonitis were observed compared to previously reported with abemaciclib and pembrolizumab monotherapy. The PK profiles were consistent between cohorts and with previous monotherapy studies. In cohorts 1/2, the overall response rate and disease control rate were 23.1/28.6% and 84.6/82.1%, respectively. Median progression-free survival and overall survivals were 8.9 (95% CI: 3.9-11.1) and 26.3 months (95% CI: 20.0-31.0) for cohort 2; cohort 1 data are immature. Abemaciclib plus pembrolizumab demonstrated antitumor activity, but high rates of ILD/pneumonitis and severe transaminase elevations occurred with/without anastrozole compared to the previous reporting. Benefit/risk analysis does not support further evaluation of this combination in the treatment of HR+, HER2- MBC.

Postantifungal Effect of Antifungal Drugs against Candida: What Do We Know and How Can We Apply This Knowledge in the Clinical Setting?

The study of the pharmacological properties of an antifungal agent integrates the drug pharmacokinetics, the fungal growth inhibition, the fungicidal effect and the postantifungal activity, laying the basis to guide optimal dosing regimen selection. The current manuscript reviews concepts regarding the postantifungal effect (PAFE) of the main classes of drugs used to treat Candida infections or candidiasis. The existence of PAFE and its magnitude are highly dependent on both the fungal species and the class of the antifungal agent. Therefore, the aim of this article was to compile the information described in the literature concerning the PAFE of polyenes, azoles and echinocandins against the Candida species of medical interest. In addition, the mechanisms involved in these phenomena, methods of study, and finally, the clinical applicability of these studies relating to the design of dosing regimens were reviewed and discussed. Additionally, different factors that could determine the variability in the PAFE were described. Most PAFE studies were conducted in vitro, and a scarcity of PAFE studies in animal models was observed. It can be stated that the echinocandins cause the most prolonged PAFE, followed by polyenes and azoles. In the case of the triazoles, it is worth noting the inconsistency found between in vitro and in vivo studies.

In vitro and in vivo anti-Candida activity of citral in combination with fluconazole.

BACKGROUND: The ability of Candida to develop biofilms on inert surfaces or living tissues favors recalcitrant and chronic candidiasis associated, in many instances, with resistance to current antifungal therapy. AIM: The aim of this study was to evaluate the antifungal activity of citral, a phytocompound present in lemongrass essential oil, in monotherapy and combined with fluconazole against azole-resistant Candida planktonic cells and biofilms. The effect of citral combined with fluconazole was also analysed with regard to the expression of fluconazole resistance-associated genes in Candida albicans and the effectiveness of the combination therapy in a Caenorhabditis elegans model of candidiasis. RESULTS: Citral reduced biofilm formation at initial stages and the metabolic activity of the mature biofilm. The combination of citral with fluconazole was synergistic, with a significant increase in the survival of C. elegans infected with Candida. RNA analysis revealed a reduction of the expression of the efflux pump encoded by MDR1, leading to a greater effect of fluconazole. CONCLUSION: Citral in monotherapy and in combination with fluconazole could represent an interesting therapy to treat recalcitrant Candida infections associated to biofilms.

Trabajando la relación Universidad-sociedad. Abordaje multidisciplinar y difusión social / Working the University-Society relationship. Multidisciplinary approach and social diffusion

INTRODUCCIÓN: La transferencia del conocimiento a la sociedad es una de las funciones importantes de la Universidad, lo que implica el uso de un lenguaje y unos medios adecuados hacia los diferentes colectivos de la sociedad, atendiendo a grupos de edad y situaciones socioeconómicas diversas. MATERIALES Y MÉTODOS: A través de las competencias transversales de cada grado, competencias genéricas que se relacionan con la puesta en práctica de una forma integrada de aptitudes, conocimientos y valores adquiridos, se ha realizado este proyecto de innovación docente con alumnado de la Universidad del País Vasco/Euskal Herriko Unibertsitatea. En él se han trabajado las habilidades del alumnado en el uso de diferentes registros de comunicación oral y escritura según la audiencia hacia la que se dirigen. Este trabajo se ha realizado dentro de un equipo multidisciplinar, de forma que el alumnado ha podido conocer y afrontar problemas de salud que requieren una actuación conjunta con otros profesionales del ámbito sanitario y científico. RESULTADOS Y CONCLUSIÓN: Esta interacción entre alumnado de diferentes grados ha permitido su enriquecimiento, proporcionándoles una visión más amplia de lo que pueden aportar los diferentes profesionales frente al mismo problema o reto. Desde este proyecto se ha planteado, a través de metodologías activas, favorecer la interacción entre los futuros profesionales de diferentes disciplinas y concienciar de la importancia de la transmisión de conocimiento a la sociedad, creando redes que contribuyan a la innovación y transferencia

INTRODUCTION: The transfer of knowledge to society is one of the important functions of the university, which implies the use of an appropriate language and means towards the different groups of society, attending to age groups and diverse socio-economic situations. MATERIALS AND METHODS: Through the transversal competences of each Degree, generic competences that are related to the implementation of an integrated form of acquired skills, knowledge and values, this teaching innovation project has been carried out with students from the University of The Basque Country (UPV / EHU). Thus, the student's abilities in the use of different oral and written communication registers according to the target audience have been studied. This work has been carried out within a multidisciplinary team, in such a way that the student has been able to know and face health problems that require joint action with other professionals in the health and scientific field. RESULTS AND CONCLUSION: This interaction among students of different degrees has allowed their enrichment, providing them with a broader vision of what different professionals can contribute to the same problem or challenge. From this project, it has been proposed, through active methodologies, to promote interaction between future professionals from different disciplines, and to raise awareness of the importance of the transmission of knowledge to society, creating networks that contribute to innovation and transfer

Disinfectant Activity of A Portable Ultraviolet C Equipment.

Healthcare-associated infections (HAIs) can be caused by microorganisms present in common practice instruments generating major health problems in the hospital environment. The aim of this work was to evaluate the disinfection capacity of a portable ultraviolet C equipment (UV Sanitizer Corvent® -UVSC-) developed to disinfect different objects. For this purpose, six pathogens causing HAIs: Acinetobacter baumannii, Bacillus subtilis, Escherichia coli, Pseudomonas aeruginosa, Staphylococcus aureus and Candida albicans, were inoculated on slides and discs of different biomaterials (borosilicate, polycarbonate, polyurethane, silicone, Teflon and titanium) and exposed to ultraviolet C radiation. UVSC disinfection was compared with ethanol and chlorhexidine antimicrobial activities following the standards EN14561 and EN14562. Disinfection, established as a reduction of five logarithms from the initial inoculum, was achieved with the UVSC at 120 s of exposure time, with and without the presence of organic matter. The disinfectant effect was observed against S. aureus, P. aeruginosa, E. coli, B. subtilis and C. albicans (reduction >99.999%). Disinfection was also achieved with 70% ethanol and 2% chlorhexidine. As conclusion, UVSC was effective disinfecting the most contaminated surfaces assayed, being a promising alternative for disinfecting hospital materials and inanimate objects that cannot be immersed in liquid biocides, reducing the risk of pathogen transmission.

Phase III evaluating the addition of fulvestrant (F) to anastrozole (A) as adjuvant therapy in postmenopausal women with hormone receptor-positive HER2-negative (HR+/HER2-) early breast cancer (EBC): results from the GEICAM/2006-10 study.

PURPOSE: GEICAM/2006-10 compared anastrozole (A) versus fulvestrant plus anastrozole (A + F) to test the hypothesis of whether a complete oestrogen blockade is superior to aromatase inhibitors alone in breast cancer patients receiving hormone adjuvant therapy. METHODS: Multicenter, open label, phase III study. HR+/HER2- EBC postmenopausal patients were randomized 1:1 to adjuvant A (5 years [year]) or A + F (A plus F 250 mg/4 weeks for 3 year followed by 2 year of A). Stratification factors: prior chemotherapy (yes/no); number of positive lymph nodes (0/1-3/≥ 4); HR status (both positive/one positive) and site. PRIMARY OBJECTIVE: disease-free survival (DFS). Planned sample size: 2852 patients. RESULTS: The study has an early stop due to the financer decision with 870 patients (437 randomized to A and 433 to A + F). Patient characteristics were well balanced. After a median follow-up of 6.24y and 111 DFS events (62 in A and 49 in A + F) the Hazard Ratio for DFS (combination vs. anastrozole) was 0.84 (95% CI 0.58-1.22; p = 0.352). The proportion of patients disease-free in arms A and A + F at 5 year and 7 year were 90.8% versus 91% and 83.6% versus 86.7%, respectively. Most relevant G2-4 toxicities (≥ 5% in either arm) with A versus A + F were joint pain (14.7%; 13.7%), fatigue (2.5%; 7.2%), bone pain (3%; 6.5%), hot flushes (3.5%; 5%) and muscle pain (2.8%; 5.1%). CONCLUSIONS: The GEICAM/2006-10 study did not show a statistically significant increase in DFS by adding adjuvant F to A, though no firm conclusions can be drawn because of the limited sample size due to the early stop of the trial. ClinicalTrials.gov: NCT00543127.

Therapeutic tools for oral candidiasis: Current and new antifungal drugs

Background: Candidiasis is one of the most common opportunistic oral infections that presents different acute and chronic clinical presentations with diverse diagnostic and therapeutic approaches. The present study carries out a bibliographic review on the therapeutic tools available against oral candidiasis and their usefulness in each clinical situation. Material and Methods: Recent studies on treatment of oral candidiasis were retrieved from PubMed and Cochrane Library. Results: Nystatin and miconazole are the most commonly used topical antifungal drugs. Both antifungal drugs are very effective but need a long time of use to eradicate the infection. The pharmacological presentations of miconazole are more comfortable for patients but this drug may interact with other drugs and this fact should be assessed before use. Other topical alternatives for oral candidiasis, such as amphotericin B or clotrimazole, are not available in many countries. Oral fluconazole is effective in treating oral candidiasis that does not respond to topical treatment. Other systemic treatment alternatives, oral or intravenous, less used are itraconazole, voriconazole or posaconazole. Available novelties include echinocandins (anidulafungin, caspofungin) and isavuconazole. Echinocandins can only be used intravenously. Isavuconazole is available for oral and intravenous use. Other hopeful alternatives are new drugs, such as ibrexafungerp, or the use of antibodies, cytokines and antimicrobial peptides. Conclusions: Nystatin, miconazole, and fluconazole are very effective for treating oral candidiasis. There are systemic alternatives for treating recalcitrant infections, such as the new triazoles, echinocandins, or lipidic presentations of amphotericin B

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Project-based learning methodology in the area of microbiology applied to undergraduate medical research.

In the recent years, there has been a decrease in the number of medical professionals dedicated to a research career. There is evidence that students with a research experience during their training acquire knowledge and skills that increase the probability of getting involved in research more successfully. In the Degree of Medicine (University of the Basque Country) the annual core subject 'Research Project' introduces students to research. The aim of this work was to implement a project-based learning methodology, with the students working on microbiology, and to analyse its result along time. Given an initial scenario, the students had to come up with a research idea related to medical microbiology and to carry out a research project, including writing a funding proposal, developing the experimental assays and analyzing and presenting their results to a congress organized by the University. Summative assessment was performed by both students and teachers. A satisfaction survey was carried out to gather the students' opinion. The overall results regarding to the classroom dynamics, learning results and motivation after the implementation were favourable. Students referred a greater interest about research than they had before. They would choose the project based methodology versus the traditional one.

Upper Tract Urothelial Carcinomas: Prognostic Factors and Outcomes in Patients With Non-Lymph Node Distant Metastasis.

BACKGROUND: Upper tract urothelial carcinomas (UTUCs) are increasingly recognized as separate malignancies. Additional insight into clinical outcomes and key prognostic factors are needed. OBJECTIVES: To detail outcomes of patients with UTUCs recurring after radical nephroureterectomy (RNU) and to determine a risk score that predicts outcomes of patients with non-lymph node distant metastasis. DESIGN, SETTING, AND PARTICIPANTS: Chart review of all patients who had an extraurothelial recurrence after RNU for UTUC at Dana-Farber Cancer Institute between 2009 and 2014. OUTCOME MEASUREMENTS AND STATISTICAL ANALYSIS: Median overall survival defined as time from chemotherapy for distant relapse to death. Prognostic relevance of performance status, hemoglobin, and receipt of cisplatin were assessed by Cox regression model. RESULTS AND LIMITATIONS: A total of 102 patients were identified, 57 of whom had non-lymph node distant metastases at relapse; 45 received chemotherapy. Median follow-up was 29.8 months; median overall survival was 14.7 months. Objective response rate to any chemotherapy in the first-line setting was only 22%. Hemoglobin > 11 g/dL and receipt of cisplatin was associated with numerically longer median survival but did not reach statistical significance in univariate and multivariate analysis. Prognostic risk score scale including hemoglobin < 11 g/dL and receipt of cisplatin was inversely associated with survival, with scores of 0, 1, and 2 leading to median survival of 19.0, 14.9, and 7.2 months (P = .38), respectively. CONCLUSIONS: Advanced UTUC portends a poor prognosis, and most patients cannot receive cisplatin-based chemotherapy. A risk score that includes anemia and receipt of cisplatin helps stratify patients with distant metastasis for inclusion into eventual clinical trials. More studies are needed to validate these findings. PATIENT SUMMARY: Metastatic UTUC is an aggressive disease, where anemia and ineligibility to receive cisplatin are adverse features associated with shorter survival.

Isolation of Pseudomonas fluorescens species highly resistant to pentachlorobenzene.

Polychlorinated aromatic compounds, including pentachlorobenzenes and hexachlorobenzenes, are recalcitrant industrial pollutants that cause adverse effects on living cells. In this paper, the isolation of Pseudomonas fluorescens species with high resistance to pentachlorobenzene (PeCB) is reported. It was found that, in contrast to its slightly negative effect on P. fluorescens growth, PeCB readily inhibited the cell growth of Serratia spp. and Escherichia coli strains, thus indicating that inhibition of bacterial growth by PeCB is species-dependent. Analysis of a P. fluorescens isolate revealed that the exposure to PeCB induced production of reactive oxygen species and led to an increase in the level of alkyl hydroperoxide reductase C (AhpC), an important enzyme enhancing the cell tolerance to organic hydroperoxides usually accumulated under oxidative stress. The putative mechanism conferring PeCB resistance to P. fluorescens and the potential use of P. fluorescens in bioremediation are discussed.

A nomogram for predicting complications in patients with solid tumours and seemingly stable febrile neutropenia.

BACKGROUND: We sought to develop and externally validate a nomogram and web-based calculator to individually predict the development of serious complications in seemingly stable adult patients with solid tumours and episodes of febrile neutropenia (FN). PATIENTS AND METHODS: The data from the FINITE study (n=1133) and University of Salamanca Hospital (USH) FN registry (n=296) were used to develop and validate this tool. The main eligibility criterion was the presence of apparent clinical stability, defined as events without acute organ dysfunction, abnormal vital signs, or major infections. Discriminatory ability was measured as the concordance index and stratification into risk groups. RESULTS: The rate of infection-related complications in the FINITE and USH series was 13.4% and 18.6%, respectively. The nomogram used the following covariates: Eastern Cooperative Group (ECOG) Performance Status ⩾2, chronic obstructive pulmonary disease, chronic cardiovascular disease, mucositis of grade ⩾2 (National Cancer Institute Common Toxicity Criteria), monocytes <200/mm(3), and stress-induced hyperglycaemia. The nomogram predictions appeared to be well calibrated in both data sets (Hosmer-Lemeshow test, P>0.1). The concordance index was 0.855 and 0.831 in each series. Risk group stratification revealed a significant distinction in the proportion of complications. With a ⩾116-point cutoff, the nomogram yielded the following prognostic indices in the USH registry validation series: 66% sensitivity, 83% specificity, 3.88 positive likelihood ratio, 48% positive predictive value, and 91% negative predictive value. CONCLUSIONS: We have developed and externally validated a nomogram and web calculator to predict serious complications that can potentially impact decision-making in patients with seemingly stable FN.

A new custom microarray for sRNA profiling in Escherichia coli.

Bacterial small RNAs (sRNAs) play essential roles in the post-transcriptional control of gene expression. To improve their detection by conventional microarrays, we designed a custom microarray containing a group of probes targeting known and some putative Escherichia coli sRNAs. To assess its potential in detection of sRNAs, RNA profiling experiments were performed with total RNA extracted from E. coli MG1655 cells exponentially grown in rich (Luria-Bertani) and minimal (M9/glucose) media. We found that many sRNAs could yield reasonably strong and statistically significant signals corresponding to nearly all sRNAs annotated in the EcoCyc database. Besides differential expression of two sRNAs (GcvB and RydB), expression of other sRNAs was less affected by the composition of the growth media. Other examples of the differentially expressed sRNAs were revealed by comparing gene expression of the wild-type strain and its isogenic mutant lacking functional poly(A) polymerase I (pcnB). Further, northern blot analysis was employed to validate these data and to assess the existence of new putative sRNAs. Our results suggest that the use of custom microarrays with improved capacities for detection of sRNAs can offer an attractive opportunity for efficient gene expression profiling of sRNAs and their target mRNAs at the whole transcriptome level.

Carbapenem-resistant and carbapenem-susceptible isogenic isolates of Klebsiella pneumoniae ST101 causing infection in a tertiary hospital.

BACKGROUND: In this study we describe the clinical and molecular characteristics of an outbreak due to carbapenem-resistant Klebsiella pneumoniae (CR-KP) producing CTX-M-15 and OXA-48 carbapenemase. Isogenic strains, carbapenem-susceptible K. pneumoniae (CS-KP) producing CTX-M-15, were also involved in the outbreak. RESULTS: From October 2010 to December 2012 a total of 62 CR-KP and 23 CS-KP were isolated from clinical samples of 42 patients (22 had resistant isolates, 14 had susceptible isolates, and 6 had both CR and CS isolates). All patients had underlying diseases and 17 of them (14 patients with CR-KP and 3 with CS-KP) had received carbapenems previously. The range of carbapenem MICs for total isolates were: imipenem: 2 to >32 µg/ml vs. <2 µg/ml; meropenem: 4 to >32 µg/ml vs. <2 µg/ml; and ertapenem: 8 to >32 µg/ml vs. <2 µg/ml. All the isolates were also resistant to gentamicin, ciprofloxacin, and cotrimoxazole. Both types of isolates shared a common PFGE pattern associated with the multilocus sequence type 101 (ST101). The bla CTX-M-15 gene was detected in all the isolates, whereas the bla OXA-48 gene was only detected in CR-KP isolates on a 70 kb plasmid. CONCLUSIONS: The clonal spread of K. pneumoniae ST101 expressing the OXA-48 and CTX-M-15 beta-lactamases was the cause of an outbreak of CR-KP infections. CTX-M-15-producing isolates lacking the bla OXA-48 gene coexisted during the outbreak.

Prediction of serious complications in patients with seemingly stable febrile neutropenia: validation of the Clinical Index of Stable Febrile Neutropenia in a prospective cohort of patients from the FINITE study.

PURPOSE: To validate a prognostic score predicting major complications in patients with solid tumors and seemingly stable episodes of febrile neutropenia (FN). The definition of clinical stability implies the absence of organ dysfunction, abnormalities in vital signs, and major infections. PATIENTS AND METHODS: We developed the Clinical Index of Stable Febrile Neutropenia (CISNE), with six explanatory variables associated with serious complications: Eastern Cooperative Oncology Group performance status ≥ 2 (2 points), chronic obstructive pulmonary disease (1 point), chronic cardiovascular disease (1 point), mucositis of grade ≥ 2 (National Cancer Institute Common Toxicity Criteria; 1 point), monocytes < 200 per µL (1 point), and stress-induced hyperglycemia (2 points). We integrated these factors into a score ranging from 0 to 8, which classifies patients into three prognostic classes: low (0 points), intermediate (1 to 2 points), and high risk (≥ 3 points). We present a multicenter validation of CISNE. RESULTS: We prospectively recruited 1,133 patients with seemingly stable FN from 25 hospitals. Complication rates in the training and validation subsets, respectively, were 1.1% and 1.1% in low-, 6.1% and 6.2% in intermediate-, and 32.5% and 36% in high-risk patients; mortality rates within each class were 0% in low-, 1.6% and 0% in intermediate-, and 4.3% and 3.1% in high-risk patients. Areas under the receiver operating characteristic curves in the validation subset were 0.652 (95% CI, 0.598 to 0.703) for Talcott, 0.721 (95% CI, 0.669 to 0.768) for Multinational Association for Supportive Care in Cancer (MASCC), and 0.868 (95% CI, 0.827 to 0.903) for CISNE (P = .002 for comparison between CISNE and MASCC). CONCLUSION: CISNE is a valid model for accurately classifying patients with cancer with seemingly stable FN episodes.

Emergence and clonal dissemination of carbapenem-hydrolysing OXA-58-producing Acinetobacter baumannii isolates in Bolivia.

Acinetobacter baumannii is an emerging multidrug-resistant pathogen and very little information is available regarding its imipenem resistance in Latin American countries such as Bolivia. This study investigated the antimicrobial resistance profile of 46 clinical strains from different hospitals in Cochabamba, Bolivia, from March 2008 to July 2009, and the presence of carbapenemases as a mechanism of resistance to imipenem. Isolates were obtained from 46 patients (one isolate per patient; 30 males,16 females) with an age range of 1 day to 84 years, and were collected from different sample types, the majority from respiratory tract infections (17) and wounds (13). Resistance to imipenem was detected in 15 isolates collected from different hospitals of the city. These isolates grouped into the same genotype, named A, and were resistant to all antibiotics tested including imipenem, with susceptibility only to colistin. Experiments to detect carbapenemases revealed the presence of the OXA-58 carbapenemase. Further analysis revealed the location of the bla(OXA-58) gene on a 40 kb plasmid. To our knowledge, this is the first report of carbapenem resistance in A. baumannii isolates from Bolivia that is conferred by the OXA-58 carbapenemase. The presence of this gene in a multidrug-resistant clone and its location within a plasmid is of great concern with regard to the spread of carbapenem-resistant A. baumannii in the hospital environment in Bolivia.

Evolution of multidrug-resistant Acinetobacter baumannii isolates obtained from elderly patients with respiratory tract infections.

OBJECTIVES: To study the evolution between 1999 and 2002 and mechanisms of antibiotic resistance in a multidrug-resistant Acinetobacter baumannii clone predominant in isolates from elderly patients with respiratory tract infections. METHODS: Susceptibility to antimicrobials was determined using an agar dilution method. Bacterial clones were identified by PCR-fingerprinting and PFGE with ApaI. Carbapenemases were detected by phenotypic tests; by PCR with primers specific for bla (OXA-40), bla(IMP), bla(VIM-1) and bla(VIM-2); and by hybridization with DNA probes. Class 1 integrons were detected using PCR. RESULTS: In 1999 isolates were grouped into two main genotypes: clone I (33%) and clone II (55%). These were also detected in 2002 with a different distribution: clone I (69%), clone II (22%). Resistance to amikacin, meropenem and imipenem increased significantly in clone I over this time, whereas clone II was not affected. In 2002, the incidence of bla(OXA-40) rose to 91% in clone I isolates with some also harbouring bla(VIM-2) and bla(IMP) genes. Different class 1 integrons were detected ranging in size from 550 to 1200 bp. No relationship was found between carbapenemases and class 1 integrons. CONCLUSIONS: In elderly patients, a single clone became predominant among A. baumannii isolates, coinciding with an increase in antibiotic resistance rates. The majority of isolates harboured the bla(OXA-40) carbapenemase gene and some of them also harboured bla(VIM-2) and bla(IMP) genes. The presence of class 1 integrons also increased over time.