Protective effects of different lyoprotectants on survival of clinical bacterial isolates in a hospital biobank.

Nowadays the significant role of biobanks in medical, diagnostic, industrial, and environmental research is well known. Bacterial biobanks could be used as a good resource for designing new treatments, biomedical and industrial researches, and laboratory diagnostics. To have a collection of bacteria from clinical samples and maintain their long-term viability, their preservation needs appropriate protective agents, like cryoprotectants and lyoprotectants. In this study, we collected and characterized Gram-negative and Gram-positive bacteria carrying important antibiotic resistance markers from different clinical samples of hospitalized children. Sucrose (10%), skimmed milk (10%), skimmed milk plus sodium glutamate (10% + 1%), and bovine serum albumin (BSA, 10%) were used as lyoprotectants during the freeze-drying procedure. The survival rate of the lyophilized samples was calculated by dilution plating and measuring the colony forming unit (CFU) after 3 months of storage. The culture analysis results indicated that 25 of the 27 studied bacterial genera (Dilutions 10-3 to 10-6), including Shigella, Methicillin-resistant S. aureus, Acinetobacter spp., Escherichia spp., Pseudomonas spp., Klebsiella spp., Enterococcus spp., were recovered in cultured fractions from all preservation conditions, while 2 genera were only detected in a single preservation condition (2/27, 7.4%). Based on the results, sucrose (10%) and skimmed milk (10%) presented the most protective features. The survival rates varied significantly according to types of the bacteria. Collectively, our results showed a diversity in the recovery of different bacterial genera after lyophilization. While statistically no significant difference was detected among the studied protective agents, sucrose (10%) and skimmed milk (10%) exhibited more effective lyoprotective properties for both Gram-positive and Gram-negative bacteria among the clinical isolates in our study.

The intestinal carrier status of Enterococcus spp. in children: clonal diversity and alterations in resistance phenotypes before and after admission to a pediatric intensive care unit.

BACKGROUND: This study aimed to investigate the intestinal carrier status of Enterococcus spp. among children in a pediatric intensive care unit (PICU) and reveal the role of hospitalization in the alteration of resistance phenotypes and clonal diversity of the isolates during admission and discharge periods. METHODS: Two separate stool samples were collected from hospitalized patients in the pediatric intensive care unit at admission and discharge times. The culture was done, and Enterococcus species were tested for antimicrobial susceptibility and carriage of vanA-D gene subtypes. Random Amplified Polymorphic DNA (RAPD)-PCR was used for a phylogenetic study to check the homology of pairs of isolates. RESULTS: The results showed carriage of Enterococci at admission, discharge, and at both time points in 31%, 28.7%, and 40.1% of the cases, respectively. High frequencies of the fecal Enterococcus isolates with vancomycin-resistance (VR, 32.6% and 41.9%), high-level of gentamicin-resistance (HLGR, 25.6% and 27.9%), and multi-drug resistance phenotypes (MDR, 48.8% and 65.1%) were detected at admission and discharge times, respectively. Resistance to vancomycin, ampicillin, and rifampicin was higher among E. faecium, but resistance to ciprofloxacin was higher in E. faecalis isolates. The increased length of hospital stay was correlated with the carriage of resistant strains to vancomycin, ampicillin, and ciprofloxacin. While the homology of the isolates was low among different patients during hospitalization, identical (9%) and similar (21%) RAPD-PCR patterns were detected between pairs of isolates from each patient. CONCLUSIONS: The high rate of intestinal carriage of VR, HLGR-, and MDR-Enterococci at admission and during hospitalization in the PICU, and the impact of increased length of hospital stay on the fecal carriage of the resistant strains show the importance of antibiotic stewardship programs to control their transmission and spread in children.

Intestinal colonization of vancomycin-resistant Enterococcus in children admitted to Mofid children's hospital intensive care unit at admission and at discharge.

BACKGROUND: This study aimed to investigate the frequency of intestinal colonization by vancomycin-resistant Enterococcus (VRE) carrying vanA and vanB genes in patients at ICU admission and at discharge from ICU in Mofid children's Hospital, Tehran, Iran. METHOD: Sampling was performed using rectal swabs and vancomycin susceptibility testing for Enterococcus spp. was carried out using a minimum inhibitory concentration (MIC) assay on Muller Hinton Agar (MHA) medium using an E-test kit. The molecular detection of VRE isolates was performed by the PCR method using the vanA and vanB resistance genes. RESULTS: A total of 234 and 186 non-duplicate rectal swab samples were collected from patients at ICU admission and at discharge from ICU, respectively. Enterococcus spp. was detected in 34.6% (n = 81/234) of rectal swab samples collected from patients at ICU admission, of which 44.4% (n = 36/81) were VRE isolates. In contrast, the prevalence of Enterococcus spp. and VRE isolates among patients at discharge from ICU was 17.7% (n = 33/186) and 57.6% (n = 19/33), respectively. Out of 19 VRE isolated from patients at ICU admission, 4 (21%) and 1 (5.3%) contained vanA and vanB genes, respectively. In contrast, out of 36 VRE isolated from patients at discharge from ICU, 11 (30.5%) were positive for the vanA gene. CONCLUSION: Results revealed that the prevalence of Enterococcus spp. among patients at ICU admission was high. However, VRE was frequently isolated from patients who were hospitalized for several days in ICUs. The implementation of proper infection control strategies and the use of suitable protocols to guide the appropriate prescribing of antibiotics are necessary.

In Vitro Antifungal Susceptibility Profiles of Candida albicans Complex Isolated from Patients with Respiratory Infections.

Candidiasis, the main opportunistic fungal infection has been increased over the past decades. This study aimed to characterize C.albicans species complex (C.albicans, C.dubliniensis, and C.africana) isolated from patients with respiratory infections by molecular tools and in vitro antifungal susceptibilities by using broth microdilution method according to CLSI M27-A3 guidelines. Totally, 121 respiratory samples were collected from patients with respiratory infections. Of these, 83 strains were germ tube positive and green colonies on chromogenic media, so initially identified as C.albicans species complex and subsequently were classified as C.albicans (89.15%), C.dubliniensis (9.63%), and C.africana (1.2%) based on PCR-RFLP and amplification of hwp1 gene. Minimum inhibitory concentration (MICs) results showed that all tested isolates of C.albicans complex were highly susceptible to triazole drugs. However, caspofungin had highest activity against C.albicans, C.dubliniensis, and C.africana. Our findings indicated the variety of antifungal resistance of Candida strains in different areas. These results may increase the knowledge about the local distribution of the mentioned strains as well as their antifungal susceptibility pattern which play an important role in appropriate therapy.