Evaluation of an in-house MALDI-TOF MS rapid diagnostic method for direct identification of micro-organisms from blood cultures.

PURPOSE: Bloodstream infections are major causes of morbidity and mortality among hospitalized patients worldwide. Early identification of micro-organisms from blood culture can facilitate earlier optimization of treatment. The objective of this study was to assess an in-house method based on a new matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) platform (Clin-TOF MS) for direct organism identification. METHODOLOGY: We studied the performance of the in-house method for direct identification and the conventional sub-culture method in parallel. Identification from subcultures was analysed with Bruker MS as the reference method. RESULTS: A total of 666 blood cultures with a single micro-organism that flagged positive after no more than a 3-day incubation period were collected. The identification accuracy of the in-house Clin-TOF MS method for direct identification and the sub-culture method was 88.6 and 100 %, respectively. The in-house method exhibited better performance for Gram-negative bacteria than for Gram-positive bacteria (93.3 vs 81.6 %). The accuracy rate for anaerobes was 100 % (3/3). The lowest accurate identification rate was for yeast; this was only 20 %. Lytic Anaerobic/F (LAF) and Plus Aerobic/F (PAF) provided the highest accurate identification rates, and it was noteworthy that the accuracy rate for FAN Aerobic (FA) was 82 %, which is higher than previously reported and showed that the method was effective. CONCLUSION: Our study provides an effective sample preparation method for the direct identification of pathogens from positive blood culture vials via Clin-TOF MS at a very low cost of about $0.5 per sample and with a short turnaround time of about 20 min. This will help clinicians make precise diagnoses and provide targeted prescriptions, reducing the risk of the potential development of resistance.

Comparative genetic characterization of Enteroaggregative Escherichia coli strains recovered from clinical and non-clinical settings.

The origin of pathogenic Enteroaggregative Escherichia coli (EAEC), a major causative agent of childhood diarrhea worldwide, remains ill-defined. The objective of this study was to determine the relative prevalence of EAEC in clinical and non-clinical sources and compare their genetic characteristics in order to identify strains that rarely and commonly cause human diarrhea. The virulence gene astA was commonly detectable in both clinical and non-clinical EAEC, while clinical isolates, but not the non-clinical strains, were consistently found to harbor other virulence factors such as aap (32%), aatA (18%) and aggR (11%). MLST analysis revealed the extremely high diversity of EAEC ST types, which can be grouped into three categories including: (i) non-clinical EAEC that rarely cause human infections; (ii) virulent strains recoverable in diarrhea patients that are also commonly found in the non-clinical sources; (iii) organisms causing human infections but rarely recoverable in the non-clinical setting. In addition, the high resistance in these EAEC isolates in particular resistance to fluoroquinolones and cephalosporins raised a huge concern for clinical EAEC infection control. The data from this study suggests that EAEC strains were diversely distributed in non-clinical and clinical setting and some of the clinical isolates may originate from the non-clinical setting.

High prevalence of qnr and aac(6')-Ib-cr genes in both water-borne environmental bacteria and clinical isolates of Citrobacter freundii in China.

We investigated the prevalence of qnr and aac(6')-Ib-cr genes in water-borne environmental bacteria and in clinical isolates of Enterobacteriaceae, as well as the subtypes of qnr. Environmental bacteria were isolated from surface water samples obtained from 10 different locations in Hangzhou City, and clinical isolates of Citrobacter freundii were isolated from several hospitals in four cities in China. qnrA, qnrB, qnrS, and aac(6')-Ib-cr genes were screened using PCR, and the genotypes were analyzed by DNA sequencing. Ten of the 78 Gram-negative bacilli isolated from water samples were C. freundii and 80% of these isolates carried the qnrB gene. qnrS1 and aac(6')-Ib-cr genes were detected in two Escherichia coli isolates and qnrS2 was detected in one species, Aeromonas punctata. The qnr and aac(6')-Ib-cr genes were present in 75 (72.8%) and 12 (11.6%) of 103 clinical isolates of C. freundii, respectively. Of the clinical C. freundii isolates with the qnr gene, 65 isolates (63.1%) carried qnrB, but only three (2.9%) and one (1.0%) carried qnrA1 and qnrS2, respectively, while five isolates carried both qnrA1 and qnrB, and one isolate carried both qnrS1 and qnrB. The qnrB9 gene was the dominant qnrB subtype, followed by qnrB8 and qnrB6. Southern hybridization studies indicated that the qnr genes are located on different plasmids. Plasmids isolated from both environmental and clinical C. freundii isolates appeared to be homogenous.

[Alendronate prevents steroid-induced osteoporosis in patients with rheumatic diseases].

OBJECTIVE: To investigate the effects of alendronate (Alen) on the prevention of systemic glucocorticoid-induced osteoporosis in patients with rheumatic diseases. METHODS: 140 patients suffering from rheumatic diseases, including systemic lupus erythematosus, polymyositis, dermatomyositis, and Sjögren's syndrome, with normal bone mineral density (BMD) and treated with oral glucocorticoids were randomly divided into 2 groups: Alen + calcium group (n = 74) receiving Alen 10 mg once a day and castrate D 600 0.6 g once a day for 24 weeks and control group (n = 66) receiving castrate D 600 0.6 g once a day for 24 weeks. The BMD and biomarkers of bone turnover were measured at baseline and 24 weeks after initiating glucocorticoid therapy. RESULTS: After 24 weeks, the BMD values at lumbar spine, femoral neck, major trochanter, and Ward' s triangle increased by 6.1%, 6.3%, 3.3%, and 2.2% respectively compared with those at baseline (all P<0.05), however, those of the control group decreased by 8.7%, 9.1%, 7.7%, and 6.4% respectively (P<0.01, P<0.05), and the BMD levels at lumbar spine and femoral neck 24 weeks later of the Alen + calcium group were both higher than those of the control group (P<0.01, P<0.05). 24 weeks later the level of urine cross linked N-telopeptides of type I collagen (NTX) of the Alen + calcium group decreased (P<0.05), and the blood osteocalcin (BGP) of the Alen + calcium group increased, however, not significantly (P>0.05). There were no significant differences in serum AKP and BGP and urine NTX 24 weeks later between these 2 groups. CONCLUSION: Improving BMD, alendronate plays an important role in the prevention of glucocorticoid-induced osteoporosis. However, calcium treatment alone fails to prevent the loss of bone.

Olfactory identification and apolipoprotein E epsilon 4 allele in mild cognitive impairment.

To investigate olfactory identification and apolipoprotein E epsilon 4 allele in patients with mild cognitive impairment (MCI), we used Cross-Cultural Smell Identification Test (CC-SIT) from University of Pennsylvania to assess olfactory identification performance and polymerase chain reaction (PCR) to detect apolipoprotein E epsilon 4 (ApoE epsilon 4) allele in 28 patients with MCI and the 30 age-matched control subjects in present study. The Mann-Whitney U test demonstrated that the MCI group performed significantly worse on CC-SIT than the normal aging group (P<0.01). For MCI patients olfaction scores correlated positively with CAMCOG-C (r=0.61, P<0.01), but not with age, gender or years of education. In normal subjects, the CC-SIT score showed no significant associations with age, gender, years of education, or CAMCOG-C. As the least common allele in Chinese, epsilon 4 was found in 13.3% of controls and in 35.8% of MCI in this study. ApoE epsilon 4 was significantly higher in MCI group than normal group (chi(2)=4.65, P<0.01). There was a significant effect of allele status on odor identification: subjects with epsilon 4 allele were not able to identify as many odors as the subjects without epsilon 4 allele (P<0.01). These results suggested that the decreased olfactory identification in MCI may be a marker for the early diagnosis of Alzheimer's disease, and ApoE genotype may be part of the basis of olfactory identification decline.

Age-associated difference in circadian sleep-wake and rest-activity rhythms.

Using actigraphic monitoring of wrist activity, we investigated the sleep and rest-activity patterns of 65 young, middle-aged, old and the oldest subjects in their natural environmental conditions. To assess the effects of age and gender on sleep and circadian rhythms in activity, multivariate analyses were performed. Age significantly affected circadian sleep and rest-activity rhythms. In the old and oldest groups, the actigraphic estimates of "actual sleep time" and "sleep efficiency" decreased significantly. The estimates of "sleep latency," the number of "nighttime awakening," sleep fragmentation and daytime naps significantly increased in the old and oldest groups. Concerning the circadian patterning of rest and activity, the interdaily stability (IS) was similar in the four age groups, while the old and oldest subjects showed significant increases in intradaily variability (IV) and nighttime activity and a decrease in amplitude (AMP). The present study demonstrated weakened and fragmented circadian sleep and rest-activity rhythms during aging. However, no gender-related difference was found.