Carbapenemase-Producing Klebsiella pneumoniae in COVID-19 Intensive Care Patients: Identification of IncL-VIM-1 Plasmid in Previously Non-Predominant Sequence Types.

During the COVID-19 pandemic, intensive care units (ICUs) operated at or above capacity, and the number of ICU patients coinfected by nosocomial microorganisms increased. Here, we characterize the population structure and resistance mechanisms of carbapenemase-producing Klebsiella pneumoniae (CP-Kpn) from COVID-19 ICU patients and compare them to pre-pandemic populations of CP-Kpn. We analyzed 84 CP-Kpn isolates obtained during the pandemic and 74 CP-Kpn isolates obtained during the pre-pandemic period (2019) by whole genome sequencing, core genome multilocus sequence typing, plasmid reconstruction, and antibiotic susceptibility tests. More CP-Kpn COVID-19 isolates produced OXA-48 (60/84, 71.4%) and VIM-1 (18/84, 21.4%) than KPC (8/84, 9.5%). Fewer pre-pandemic CP-Kpn isolates produced VIM-1 (7/74, 9.5%). Cefiderocol (97.3-100%) and plazomicin (97.5-100%) had the highest antibiotic activity against pandemic and pre-pandemic isolates. Sequence type 307 (ST307) was the most widely distributed ST in both groups. VIM-1-producing isolates belonging to ST307, ST17, ST321 and ST485, (STs infrequently associated to VIM-1) were detected during the COVID-19 period. Class 1 integron Int1-blaVIM-1-aac(6')-1b-dfrB1-aadAI-catB2-qacEΔ1/sul1, found on an IncL plasmid of approximately 70,000 bp, carried blaVIM-1 in ST307, ST17, ST485, and ST321 isolates. Thus, CP-Kpn populations from pandemic and pre-pandemic periods have similarities. However, VIM-1 isolates associated with atypical STs increased during the pandemic, which warrants additional monitoring and surveillance.

Legionella feeleii: Ubiquitous Pathogen in the Environment and Causative Agent of Pneumonia.

L. feeleii is one of the most frequent Legionella species isolated from natural pools of the central region of Spain. This study aimed to evaluate its ecology and to identify this Legionella species as a respiratory pathogen. A PCR assay for detecting the L. feeleii mip gene was developed to identify it in clinical and environmental samples. Culture and PCR were performed in environmental samples from four drinking water treatment plants (DWTPs). Free L. feeleii was only detected in raw water samples (3.4%), while L. feeleii as an Acanthamoeba endosymbiont was found in 30.7% of raw water, 11.5% of decanter biofilm, and 32% of finished water samples. Therefore, Acanthamoeba spp. plays an essential role in the multiplication, persistence, and spread of Legionella species in the environment. The first case of Legionnaires' disease caused by L. feeleii in Spain is described in this study. The case was diagnosed in an older woman through PCR and sequencing from urine and sputum samples. A respiratory infection could be linked with health care procedures, and the patient presented several risk factors (age, insulin-dependent diabetes, and heart disease). The detection of non-L. pneumophila, such as L. feeleii, is a factor that must be considered when establishing or reviewing measures for the control and prevention of legionellosis.

Characterization of Carbapenemase-Producing Klebsiella oxytoca in Spain, 2016-2017.

There is little information about carbapenemase-producing (CP) Klebsiella oxytoca, an important nosocomial pathogen. We characterized CP K. oxytoca isolates collected from different Spanish hospitals between January 2016 and October 2017. During the study period, 139 nonduplicate CP K. oxytoca isolates were identified; of these, 80 were studied in detail. Carbapenemase and extended-spectrum ß-lactamase genes were identified by PCR and sequencing. Genetic relatedness was studied by pulsed-field gel electrophoresis (PFGE). Whole-genome sequencing (WGS), carried out on 12 representative isolates, was used to identify the resistome, to elucidate the phylogeny, and to determine the plasmids harboring carbapenemase genes. Forty-eight (60%) isolates produced VIM-1, 30 (37.5%) produced OXA-48, 3 (3.7%) produced KPC-2, 2 (2.5%) produced KPC-3, and 1 (1.2%) produced NDM-1; 4 isolates coproduced two carbapenemases. By PFGE, 69 patterns were obtained from the 80 CP K. oxytoca isolates, and four well-defined clusters were detected: cluster 1 consisted of 11 OXA-48-producing isolates, and the other three clusters included VIM-1-producing isolates (5, 3, and 3 isolates, respectively). In the 12 sequenced isolates, the average number of acquired resistance genes was significantly higher in VIM-1-producing isolates (10.8) than in OXA-48-producing isolates (2.3). All 12 isolates had chromosomally encoded genes of the blaOXY-2 genotype, and by multilocus sequence typing, most belonged to sequence type 2 (ST2). Carbapenemase genes were carried by IncL, IncHI2, IncFII, IncN, IncC, and IncP6 plasmid types. The emergence of CP K. oxytoca was principally due to the spread of VIM-1- and OXA-48-producing isolates in which VIM-1- and OXA-48 were carried by IncL, IncHI2, IncFII, and IncN plasmids. ST2 and the genotype blaOXY-2 predominated among the 12 sequenced isolates.

Correction: First Case of Legionnaire's Disease Caused by Legionella anisa in Spain and the Limitations on the Diagnosis of Legionella non-pneumophila Infections.

[This corrects the article DOI: 10.1371/journal.pone.0159726.].

First Case of Legionnaire's Disease Caused by Legionella anisa in Spain and the Limitations on the Diagnosis of Legionella non-pneumophila Infections.

Legionnaires' disease is a severe form of pneumonia, with worldwide relevance, caused by Legionella spp. Approximately 90% of all cases of legionellosis are caused by Legionella pneumophila, but other species can also be responsible for this infection. These bacteria are transmitted by inhalation of aerosols or aspiration of contaminated water. In Spain, environmental studies have demonstrated the presence of Legionella non-pneumophila species in drinking water treatment plants and water distribution networks. Aware that this evidence indicates a risk factor and the lack of routine assays designed to detect simultaneously diverse Legionella species, we analyzed 210 urine samples from patients presenting clinical manifestations of pneumonia using a semi-nested PCR for partial amplification of the 16S rDNA gene of Legionella and a diagnostic method used in hospitals for Legionella antigen detection. In this study, we detected a total of 15 cases of legionellosis (7.1%) and the first case of Legionnaires' disease caused by L. anisa in Spain. While the conventional method used in hospitals could only detect four cases (1.9%) produced by L. pneumophila serogroup 1, using PCR, the following species were identified: Legionella spp. (10/15), L. pneumophila (4/15) and L. anisa (1/15). These results suggest the need to change hospital diagnostic strategies regarding the identification of Legionella species associated with this disease. Therefore, the detection of Legionella DNA by PCR in urine samples seems to be a suitable alternative method for a sensitive, accurate and rapid diagnosis of Legionella pneumonia, caused by L. pneumophila and also for L. non-pneumophila species.

[Epidemiology of invasive candidiasis in pediatric and adult populations]. / Epidemiología de las candidiasis invasoras en población pediátrica y adulta.

BACKGROUND: Invasive candidiasis (IC) is the most frequent fungal disease in children and adults. AIMS: To critically review and update the current epidemiology of Candida spp. disease in neonates, children and adults (critically ill patients and in oncohematologic patients and in solid organ transplant recipients). METHODS: We searched the PubMed/Medline, discussing the current data. RESULTS AND CONCLUSIONS: IC is associated with high attributable morbimortality and increased healthcare costs. In the last decades the incidence of invasive Candida spp. disease has increased in critically ill patients, has decreased in oncohematologic patients, although currently the involvement of non-albicans Candida species in the etiology of this disease is increasing steadily.

Detection of (1-->3)-beta-D-glucan as an adjunct to diagnosis in a mixed population with uncommon proven invasive fungal diseases or with an unusual clinical presentation.

This single-center observational prospective study evaluated the performance of (1-->3)-beta-D-glucan as an adjunct diagnostic tool in 12 patients with proven invasive fungal disease with different risk factors. The infections were due to either uncommon fungal pathogens such as dematiaceous molds (Scedosporium apiospermum, Alternaria infectoria, and Cladosporium macrocarpum) and hyaline septate molds (Fusarium solani and Blastoschizomyces capitatus) or Aspergillus spp. with unusual clinical presentations.

Isolation of Aspergillus lentulus in Spain from a critically ill patient with chronic obstructive pulmonary disease.

Aspergillus lentulus was first described in the year 2005, and since it cannot be phenotypically distinguished from Aspergillus fumigatus, it is conceivable that earlier descriptions (before 2005) could be attributed to this new species. Currently invasive infections caused by A. lentulus are rare and very few cases have been previously published in neutropenic patients, all of them with fatal outcome. Here we report a critically ill non neutropenic patient with chronic obstructive pulmonary disease (COPD) who was admitted to the medical intensive care unit with an exacerbation of COPD and who had been treated with long term corticosteroids. A. fumigatus was cultured from two bronchial aspirates and in a third bronchial aspirate both A. lentulus and A. fumigatus were isolated. On two consecutive days detection of galactomannan in serum was negative whilst detection of (1-3) beta-D glucan was positive (> 518 pg/ml). Minimal inhibitory concentrations (MIC) for itraconazole, voriconazole, caspofungin and amphotericin B were high for this strain of A. lentulus. Given the high MIC values of A. lentulus to available antifungals, the accurate identification of this new species is warranted. To our knowledge, this is the first report of the isolation of A. lentulus in a non-neutropenic critically ill patient, although we note that since it was isolated only once from respiratory specimens, its implication as an etiologic agent of infection for this patient remains to be established.

Rhinocerebral zygomycosis caused by Saksenae vasiformis in a diabetic patient.

We report a rare case of invasive rhinocerebral Saksenae vasiformis infection in a 71-year-old man with type 2 diabetes mellitus ketoacidosis. Combined histological and microbiological examination of two biopsy specimens taken at four days' intervals showed broad non-septate hyphae on direct smears of biopsies and necrosis and angioinvasion by hyphae were seen in the tissue. S. vasiformis was cultured from both biopsies. The patient had a fatal outcome even though partial debridement, liposomal amphotericin B treatment and control of diabetes were instituted. Permission for necropsy was denied. Clinical, epidemiological and pathological features of the case and manifestations of S. vasiformis infections are discussed. Most cases involve cutaneous sites and only two cases of rhinocerebral infections (both with fatal outcome) have been described. The present case is the third documented case of involvement of the sinuses and the second case of S. vasiformis infection documented in Europe.

[Early diagnosis of invasive fungal infections caused by Aspergillus and other emerging mycelial fungi]. / Estado actual del diagnóstico precoz de las infecciones invasoras causadas por Aspergillus y otros hongos filamentosos emergentes.

The usefulness of surrogate markers in the diagnosis of invasive fungal infections caused by Aspergillus and other emerging mycelial fungi is based on the ability of surrogate markers to detect the infection caused by different species of mycelial fungi. Conventional microbiological methods for diagnosis of fungal disease are slow and insensitive. Antigen based assays or measurement of (1-3)-beta-D-glucan in blood have been developed and validated in clinical laboratories. We review these diagnostic contemporary tools, their clinical application and impact.

False positive galactomannan results in adult hematological patients treated with piperacillin-tazobactam.

In this prospective study including 78 adult patients with haematological malignancy (90 episodes) we performed galactomannan (GM) (Platelia Aspergillus) screening twice weekly for the diagnosis of invasive aspergillosis. There were five proven and four probable invasive aspergillosis cases. The sensitivity, specificity and positive and negative predictive values were 100, 88, 47 and 100%, respectively. There were eight patients with false positive GM (10.2%). In six patients the false GM reactivity was due to the administration of piperacillin-tazobactam (P-T). A significant association was found between false positive GM (= or > 0.5) and the administration of P-T (p < 0.01). Two other patients with no invasive aspergillosis (2.5%) and false GM reactivity had graft versus host disease (GVHD) and one of them had also mucositis grade IV. The kinetic patterns of false positive GM due to P-T is discussed.

[Traditional microbiological diagnosis for invasive candidiasis in critical non-neutropenic patients]. / Diagnóstico microbiológico tradicional de la candidiasis invasora en el enfermo crítico no neutropénico.

Invasive candidiasis is the most prevalent fungal infection in the critical non neutropenic patient (80%) and is associated with high morbidity-mortality. Microbiological diagnosis is difficult and the positivity of traditional tests appears late in the course of infection. We herein discuss the utility of direct examination and cultures from different sites and the value of surveillance cultures for establishing the likelihood of invasive candidiasis.

[Risk factors in invasive candidiasis: stratification]. / Factores de riesgo de la candidiasis invasora: estratificación.

Risk factors of invasive candidiasis in the setting of non neutropenic critical patients are well known, although currently there is a need to define and validate in prospective multicenter studies risk assessment strategies that would predict accurately the likelihood of invasive candidiasis. The clinical application in order to define which patients should be treated with antifungal prophylaxis and which groups or subgroups of patients should be assessed prospectively during the risk period in order to validate the new diagnostic microbiological indirect techniques for invasive candidiasis and preemptive treatment should be based in these strategies.

[Management strategies: prophylaxis, empirical and preemptive treatment of established invasive candidiasis in the critically ill non-neutropenic patient]. / Estrategias de tratamiento: profilaxis, tratamiento empírico, precoz (anticipado) y dirigido de candidiasis invasora en el enfermo crítico no neutropénico.

In critically ill non neutropenic patients there are four broad approaches for the management of antifungal treatment for invasive candidiasis: prophylaxis, empirical, preemptive therapy and treatment of established infections. All these approaches in relationship with risk strategies and microbiological indirect laboratory techniques for establishing invasive candidiasis will be discussed.

Isolation of Issatchenkia occidentalis from the esophagus of a leukemic patient.

Issatchenkia occidentalis was isolated from an esophageal biopsy of a young leukemic male patient who underwent bone marrow transplantation. At the time the specimen was collected, the patient was also suffering from esophageal herpetic lesions. The identification of the isolate was not possible by the use of the available commercial methods. Thus, its identification was done by PCR and DNA sequencing using panfungal primers.

[Activity of ertapenem and 19 other antimicrobial agents against beta-lactam-resistant Streptococcus pneumoniae and Haemophilus influenzae respiratory tract isolates]. / Actividad de ertapenem y otros 19 antimicrobianos frente a Streptococcus pneumoniae y Haemophilus influenzae aislados del tracto respiratorio resistentes a betalactámicos.

INTRODUCTION: Ertapenem is a new carbapenem with a long half-life that has a broad spectrum of action against Gram-positive and Gram-negative bacteria. Although, clinically indicated for the treatment of the polymicrobial abdominal infection, other uses are possible, due to the broad spectrum of action of this antimicrobial agent. OBJECTIVE: The purpose of this study was to examine the susceptibility to ertapenem of Streptococcus pneumoniae and Haemophilus influenzae strains with resistance to beta-lactam antibiotics. METHODS: The study comprised 105 S. pneumoniae strains with a reduced susceptibility to penicillin G, 86 S. pneumoniae strains resistant to penicillin, and 100 H. influenzae strains resistant to ampicillin isolated fron clinical respiratory samples collected in the 21 participating microbiology laboratories. Susceptibility to antimicrobials was tested using an agar dilution method. RESULTS: The S. pneumoniae strains with intermediate resistance to penicillin were highly susceptible to carbapenems, with MIC90 of 0,25 and 0,5 mg/l for imipenem and ertapenem, respectively. Among the penicillin-resistant strains the MIC90 values for imipenem and ertapenem were 1 and 2 mg/l respectively. Carbapenems were very active against the whole group of H. influenzae strains, particullary ertapenem, which inhibited 90% of the series at a concentration of 0,12 mg/l. CONCLUSIONS: Ertapenem is a powerful agent against the most common respiratory bacterial pathogens, including those with resistance mechanisms effective against other compouunds. Few strains can currently be considered resistant to this carbapenem, which is at least comparable to imipenem, cefotaxime or the modern fluorquinolones. Such performance, and its favorable pharmacokinetic characteristics, make ertapenem an interesting option for the treatment of respiratory tract infections.