Multicenter study of epidemiological cutoff values and detection of resistance in Candida spp. to anidulafungin, caspofungin, and micafungin using the Sensititre YeastOne colorimetric method.

Neither breakpoints (BPs) nor epidemiological cutoff values (ECVs) have been established for Candida spp. with anidulafungin, caspofungin, and micafungin when using the Sensititre YeastOne (SYO) broth dilution colorimetric method. In addition, reference caspofungin MICs have so far proven to be unreliable. Candida species wild-type (WT) MIC distributions (for microorganisms in a species/drug combination with no detectable phenotypic resistance) were established for 6,007 Candida albicans, 186 C. dubliniensis, 3,188 C. glabrata complex, 119 C. guilliermondii, 493 C. krusei, 205 C. lusitaniae, 3,136 C. parapsilosis complex, and 1,016 C. tropicalis isolates. SYO MIC data gathered from 38 laboratories in Australia, Canada, Europe, Mexico, New Zealand, South Africa, and the United States were pooled to statistically define SYO ECVs. ECVs for anidulafungin, caspofungin, and micafungin encompassing ≥97.5% of the statistically modeled population were, respectively, 0.12, 0.25, and 0.06 µg/ml for C. albicans, 0.12, 0.25, and 0.03 µg/ml for C. glabrata complex, 4, 2, and 4 µg/ml for C. parapsilosis complex, 0.5, 0.25, and 0.06 µg/ml for C. tropicalis, 0.25, 1, and 0.25 µg/ml for C. krusei, 0.25, 1, and 0.12 µg/ml for C. lusitaniae, 4, 2, and 2 µg/ml for C. guilliermondii, and 0.25, 0.25, and 0.12 µg/ml for C. dubliniensis. Species-specific SYO ECVs for anidulafungin, caspofungin, and micafungin correctly classified 72 (88.9%), 74 (91.4%), 76 (93.8%), respectively, of 81 Candida isolates with identified fks mutations. SYO ECVs may aid in detecting non-WT isolates with reduced susceptibility to anidulafungin, micafungin, and especially caspofungin, since testing the susceptibilities of Candida spp. to caspofungin by reference methodologies is not recommended.

Comparative evaluation of Vitek 2 identification and susceptibility testing of urinary tract pathogens directly and isolated from chromogenic media.

We evaluated the use of urine specimens for direct identification and antibiotic testing of urinary tract pathogens using the Vitek system. A total of 343 urine specimens from patients with suspected UTI were selected by pyuria and screened by Gram staining to detect bacteriuria. Of those, 132 were analysed after Gram staining, showing a high number of micro-organisms of a single morphological type. Direct susceptibility testing and identification were performed by using the Vitek system. Results were compared using the standard inoculation method based on the incubation of solid media. After sub-culture, 107 specimens grew a significant count of a single species and were used for the comparative analysis. The direct method correctly identified 88 isolates (82.3 %). When compared according to antibiotic susceptibility testing, the error rate was 2.4 % overall with 0.2 % very major, 0.4 % major and 1.8 % minor errors. 84.7 % of the Gram-negative bacilli had a complete susceptibility report in ≤ 8 h. This method offers the advantage of prompt processing and earlier reporting of complete results for positive urine specimens.

Comparative evaluation of Vitek 2 identification and susceptibility testing of Gram-negative rods directly and isolated from BacT/ALERT-positive blood culture bottles.

The performance of Vitek 2 was evaluated for the identification and susceptibility testing of Gram-negative bacilli directly from positive blood cultures bottles. Direct inoculation of the positive blood cultures with the Vitek cards ID-GN and AST-NO58 was compared with the standard inoculation method based on the sub-culture of the positive blood culture to agar. A total of 142 blood cultures were included in the study; of those, 119 were from patients' clinical samples, while 23 were artificially prepared with strains showing different mechanisms of resistance. A total of 136 (95.8%) strains were correctly identified to the species level, only 2 (1.4%) were mis-identified and 4 (2.8%) were not identified. Susceptibility results were available for all isolates tested against 17 antibiotics, thus, resulting in a total of 2,414 isolate/anti-microbial combinations. The error rate was 2.8% (67/2,414) overall; 0.6% (14/2,414) very major errors, 0.1% (3/2,414) major errors and 2.1% (50/2,414) minor errors. The direct method detected 88.5% (22/25) of the strains producing extended-spectrum beta-lactamases (ESBLs). The susceptibility agreement among the added strains with ESBL, AMPc hyperproduction, resistance to ceftazidime, carbapenems and cefepime was very high. Direct identification and susceptibility testing gave rapid and reliable results, reducing by 24 h the turnaround time of the microbiology laboratory.

Evaluation of three immunochromatographic assays for detection of Legionella pneumophila serogroup 1 antigen in urine samples.

The Uni-Gold, the SAS and the Binax NOW immunochromatographic test (ICT) urinary antigen assays for the qualitative detection of Legionella pneumophila serogroup 1 were compared using 39 unfrozen and nonconcentrated urine samples from patients with Legionnaires disease (LD). The Uni-Gold antigen test detected the urinary antigen in 41% (16/39), the SAS antigen test in 61.5% (24/39), and the Binax NOW antigen test in 74.3% (29/39). The Binax NOW ICT assay showed the best results when detecting L. pneumophila urinary antigen.

A genetic fusion GDNF-C fragment of tetanus toxin prolongs survival in a symptomatic mouse ALS model.

PURPOSE: Amyotrophic Lateral Sclerosis (ALS) is a paralyzing disorder that kills individuals within three to five years of onset without any possibility for effective treatment. One proposed therapy has been the use of neurotrophic factors to inhibit the apoptosis of motorneurones. At the present, one way to deliver neurotrophic factors after intramuscular injection to the motor neurones is through the use of adenoviral vectors. An alternative strategy is the use of the atoxic C fragment of tetanus toxin (TTC) as a neurotrophic factor carrier for motorneurones. METHODS: We have produced the recombinant protein fusion Glial Derived Neurotrophic Factor and C fragment of tetanus toxin (GDNF-TTC) and we have tested its antiapoptotic activity in degeneration culture cells and in the symptomatic SOD;{G93A} transgenic animal model for ALS. RESULTS: We demonstrated that GDNF-TTC induces the neuronal survival Akt kinase pathway in mouse cortical culture neurons and~maintains its antiapoptotic neuronal activity in Neuro2A cells. Moreover, we have found that genetic fusion is able to increase survival by 9 days and improves life quality in symptomatic ALS animal models. CONCLUSION: These results suggest that recombinant GDNF-TTC fusion protein intramuscular injections provide a potential therapy for ALS treatment.

[Nosocomial infections caused by multiresistant Pseudomonas aeruginosa (carbapenems included): predictive and prognostic factors. A prospective study (2016-2017))]. / Infecciones nosocomiales por Pseudomonas aeruginosa multiresistente incluido carbapenémicos: factores predictivos y pronósticos. Estudio prospectivo 2016-2017.

OBJECTIVE: Pseudomonas aeruginosa is one of the major pathogens causing hospital-acquired infections. In recent years, antimicrobial resistance is increasing and multidrug resistant (MDR) and extremely drug resistant (XDR) isolates have been associated with an increase of mortality. The aim of this study is to assess the clinical significance and analyze predictors and prognostic factors. METHODS: Prospective case-control non-paired study involving 64 patients with P. aeruginosa nosocomial infection, 32 caused by susceptible P. aeruginosa and 32 by MDR/XDR including to carbapenems (XDR-C) strains, admitted at a third level hospital. The follow-up period was till hospital discharge or death and at 30 days after discharge. For all patients, clinical epidemiology and microbiological data were analyzed. RESULTS: The incidence of MDR/XDR-C strains was 2.3 per 1000 admissions. Ten of which were VIM metallo-ß-lactamase-producing. Independent predictor factors associated with MDR/XDR-C infections were: previous ICU or Resuscitation unit admission (OR 14.01; IC 95% 2.105-93.297) appearance >20 days after admission (OR 29.826; IC 95% 4.783-185.997) and leukocytosis (OR 10.0190; IC 95% 1.842-56.369). However, there were not statistically significant differences in clinical severity or mortality between both groups. CONCLUSIONS: The major risk factors associated with MDR/XDR-C infections were previous ICU or Resuscitation unit admission, appearance >20 days after admission and leukocytosis. MDR/XDR-C infections were not associated to increased mortality.

Clinical Significance of Contamination of the Preservation Solution in Liver Transplantation.

INTRODUCTION: The aim of the present study was to describe the incidence and microbiological profiles of positive cultures obtained from preservation solution (PS) and correlate these findings with infectious complications detected in the liver transplant (LT) recipient. PATIENTS: We conducted a single-center, retrospective study between December 2010 and August 2014 among 178 LT. In all grafts, a PS culture was carried out. All the infections in the receipt until hospital discharge were collected. In patients with >1, infection was considered the most severe according to Clavien-Dindo classification. RESULTS: PS culture was positive for bacterial or fungal agents in 79 of 178 LT recipients (44%). The most commonly cultured organisms were coagulase-negative staphylococci (64%), Enterobacteriaceae (17%), and Staphylococcus aureus (4.7%). In the 79 patients with positive PS, 49 blood cultures were requested in the period after LT. Twenty-five postoperative infections (31.7%) were diagnosed. Only 4 of 79 patients (5%) with PS contamination had a postoperative infections related with isolated microorganism. CONCLUSIONS: Contamination of PS appears in a high percentage of liver grafts before LT, although there is a poor correlation with postoperative infections in LT recipient. In these patients, a standardized process including fungal and bacterial cultures could be useful.

Acyl phosphatidylglycerol: a major phospholipid of Corynebacterium amycolatum.

The type strain and several clinical isolates of Corynebacterium amycolatum were examined for lipid composition as a chemotaxonomic character for routine identification. The phospholipid profile was composed of diphosphatidylglycerol, phosphatidylglycerol, phosphatidylinositol and phosphatidylinositol mannosides, together with various unidentified compounds. One of them, accounting for 20-29% of total phospholipids, was purified and characterized as acyl phosphatidylglycerol by chromatographic and spectrometric techniques. The acyl substituents on the phosphatidyl moiety were characterized as tetradecanoyl, pentadecanoyl, hexadecenoyl, hexadecanoyl, heptadecenoyl, heptadecanoyl, octadecenoyl (the major one), and octadecanoyl. The acyl group on the polar head (glycerol) was only octadecenoyl. Phospholipid analysis by thin-layer chromatography of a collection of Corynebacterium strains proved that this compound is widely distributed, although it only represents a minor (2-9%) component among mycolic acid-containing species. Acyl phosphatidylglycerol can be considered as a useful chemical marker for the identification of C. amycolatum in addition to the absence of mycolic acids.

[Multidrug resistant Acinetobacter baumanii:clinical update and new highlights]. / Acinetobacter baumanii multirresistente:situación clínica actual y nuevas perspectivas.

The role of multidrug resistant Acinetobacter baumanii and its clinical relevance have been recently appreciated as a ubiquitous opportunistic nosocomial pathogen. Risk factors associated with A. baumanii infection include severe underlying diseases, previous surgery, invasive procedures, treatment with broad-spectrum antibiotics, length of hospital stay, admission to intensive care units (ICU). Carbapenem-multidrug resistant A. baumanii infections are probably associated to greater severity and more complications; in our cohort mortality was 49.3% and related mortality (within 72 hours) was 10.39%. However, severe underlying diseases probably play an important role in the clinical outcome of patients with MDR-C A. baumanii infection and controversy exists regarding the real mortality attributable to antimicrobial resistance because a high proportion of deaths took place > 7 days after diagnosis. Nevertheless, in our experience, carbapenem resistance, inappropriate therapy and monotherapy are associated to a higher mortality. Special attention should be paid to design well-controlled prospective clinical trials to determine the optimal antimicrobial therapy in critically ill patients suspected of having MDR Acinetobacter infection.

Bayesian estimates of genetic parameters for pre-conception traits, gestation length and calving interval in beef cattle.

A total of 5253 records obtained from 2081 Rubia Gallega beef cows managed using artificial insemination as the only reproduction system were analysed to estimate genetic parameters for days to first insemination (DFI), days from first insemination to conception (FIC), number of inseminations per conception (IN), days open (DO), gestation length (GL) and calving interval (CI) via multitrait Bayesian procedures. Estimates of the mean of posterior distribution of the heritability of DFI, FIC, IN, DO, GL and CI were, respectively, 0.050, 0.078, 0.071, 0.053, 0.037 and 0.085 and the corresponding estimates for repeatability of these traits were 0.116, 0.129, 0.147, 0.138, 0.082 and 0.132, respectively. No significant genetic correlations associated to DFI or GL were found. However, genetic correlations between the other four analysed traits were high and significant. Genetic correlations between FIC and IN, DO and CI were similar and higher than 0.85. Genetic correlations of IN-DO and IN-CI were over 0.65. The highest genetic correlation was estimated for the pair DO-CI (0.992) that can be considered the same trait in genetic terms. Results indicated that DFI can be highly affected by non-genetic factors thus limiting its usefulness to be used as an earlier indicator of reproductive performance in beef cattle. Moreover, GL could not be associated to the reproductive performance of the cow before conception. The other four analysed traits, FIC, IN, DO and CI, have close genetic relationships. The inclusion of IN as an earlier indicator of fertility in beef cattle improvement programs using artificial insemination as the main reproductive system can be advisable due to the low additional recording effort needed.

Pneumonia due to Nocardia cyriacigeorgica in a patient with Crohn's disease treated with infliximab.

Infliximab, an anti-tumor necrosis factor α (TNF-α) antibody, is useful in the treatment of rheumatoid arthritis, Crohn's disease etc. It has been related to increases in the rate of several infections. We present the case of a 53-year-old woman diagnosed with community-acquired pneumonia due to Nocardia cyriacigeorgica who was taking infliximab, azathioprine and prednisone for Crohn's disease.

Detection of mycoloylglycerol by thin-layer chromatography as a tool for the rapid inclusion of corynebacteria of clinical origin in the genus Corynebacterium.

A chemotaxonomic study of some corynebacteria isolated from clinical samples revealed characteristic thin-layer chromatographic patterns for meso-diaminopimelic acid containing species included in the genera Corynebacterium, Dermabacter and Brevibacterium. Notably, a specific compound was consistently detected in mycolic acid containing species of the genus Corynebacterium. This compound was composed by glycerol and mycolic acids and structural analyses carried out by fast atom bombardment mass spectrometry in C. minutissimum confirmed its identification as mycoloylglycerol. The chain length of mycoloyl groups in this molecule ranged from 28 to 34 carbon atoms, being mono-, di- or triunsaturated. Detection of mycoloylglycerol by thin-layer chromatography may be thus useful for the rapid inclusion of a great variety of corynebacteria of clinical origin in the genus Corynebacterium in laboratories employing chromatographic techniques as an adjunct for the identification of these microorganisms.

Quinolone resistance mutations in Streptococcus pneumoniae GyrA and ParC proteins: mechanistic insights into quinolone action from enzymatic analysis, intracellular levels, and phenotypes of wild-type and mutant proteins.

Mutations in DNA gyrase and/or topoisomerase IV genes are frequently encountered in quinolone-resistant mutants of Streptococcus pneumoniae. To investigate the mechanism of their effects at the molecular and cellular levels, we have used an Escherichia coli system to overexpress S. pneumoniae gyrase gyrA and topoisomerase IV parC genes encoding respective Ser81Phe and Ser79Phe mutations, two changes widely associated with quinolone resistance. Nickel chelate chromatography yielded highly purified mutant His-tagged proteins that, in the presence of the corresponding GyrB and ParE subunits, reconstituted gyrase and topoisomerase IV complexes with wild-type specific activities. In enzyme inhibition or DNA cleavage assays, these mutant enzyme complexes were at least 8- to 16-fold less responsive to both sparfloxacin and ciprofloxacin. The ciprofloxacin-resistant (Cip(r)) phenotype was silent in a sparfloxacin-resistant (Spx(r)) S. pneumoniae gyrA (Ser81Phe) strain expressing a demonstrably wild-type topoisomerase IV, whereas Spx(r) was silent in a Cip(r) parC (Ser79Phe) strain. These epistatic effects provide strong support for a model in which quinolones kill S. pneumoniae by acting not as enzyme inhibitors but as cellular poisons, with sparfloxacin killing preferentially through gyrase and ciprofloxacin through topoisomerase IV. By immunoblotting using subunit-specific antisera, intracellular GyrA/GyrB levels were a modest threefold higher than those of ParC/ParE, most likely insufficient to allow selective drug action by counterbalancing the 20- to 40-fold preference for cleavable-complex formation through topoisomerase IV observed in vitro. To reconcile these results, we suggest that drug-dependent differences in the efficiency by which ternary complexes are formed, processed, or repaired in S. pneumoniae may be key factors determining the killing pathway.

Assessment of multiple coagulase-negative staphylococci isolated in blood cultures using pulsed-field gel electrophoresis.

One of the criteria used to determine the clinical importance of coagulase-negative staphylococci (CNS) is the isolation of the bacteria from sequential blood cultures. Sequential isolates of CNS obtained from five immunocompromised patients over three months were genetically characterized by pulsed-field gel electrophoresis (PFGE). This typing method was compared to two first-line typing methods: determination of the species and of antibiotic susceptibility. In four patients the initial clinical evaluation changed because of the PFGE results several episodes of bacteremia would have been wrongly assessed if only the biotype and the antibiotype had been determined. Pulsed-field gel electrophoresis should therefore be used for CNS strains from immunocompromised patients or those suffering from chronic diseases with non-concordant biotype and anti-biotype.