Evaluation of Four Blood Culture Systems Using Parallel Culture Methods

Commercially prepared Vacutainer thioglycolate broth with SPS (TBSPS), the B-D Vacutainer culture tube with SPS (VCT), the Pfizer E-Vac hypertonic brucella broth with SPS (BBSPS), and our own Trypticase-soy hypertonic broth with SPS (TSBH) were studied by parallel culture methods for their ability to allow the survival of a wide spectrum of organisms, their ability to allow recovery of organisms, and the time required to obtain a recovery from clinical blood cultures. Organisms considered to be clinically significant were recovered from 176 blood cultures. The TSBH system produced all 176 recoveries, the BBSPS system allowed for 170 recoveries, the TBSPS recovered 131, and the VCT accounted for 106 recoveries. The time required to recover any given organism from both the BBSPS and the TSBH systems did not exceed 48 h, whereas 86 of the 131 recoveries from the TBSPS system required more than 72 h and 74 of the 106 VCT recoveries also required more than 72 h. A wide spectrum of organisms were recovered from both the BBSPS and the TSBH systems. The TBSPS system failed to allow the recovery of many streptococci and Pseudomonas spp. and the VCT system failed to recover many organisms.     

Bacteremia After Tooth Extractions Studied with the Aid of Prereduced Anaerobically Sterilized Culture Media

Both prereduced molten agar and broth and aerobic molten agar and broth were inoculated with blood samples collected from patients with periodontitis, but in otherwise good health, both before and after extraction of two or more teeth. Postoperative blood samples from 23 of 25 patients sampled yielded anaerobic and facultative species. Colony counts from nine samples yielded from less than 1 to over 100 colonies per ml of blood. Organisms detected were species belonging to the genera Bacteroides, Fusobacterium, Peptostreptococcus, Leptotrichia, Propionibacterium, Peptococcus, Veillonella, plus Streptococcus mitis, S. salivarius, vibrio forms, and strains resembling S. mutans. The data indicate that prereduced anaerobically sterilized culture medium with polyanethol sulfonate is effective for detecting anaerobic species in bacteremia and that anaerobic species can be prevalent in bacteremias immediately after tooth extraction in patients with periodontitis.     

Growth of Leptospira pomona and Its Effect on Various Tissue Culture Systems

Miller, Robert E. (University of Nebraska College of Medicine, Omaha), Norman G. Miller, and Roberta J. White. Growth of Leptospira pomona and its effect on various tissue culture systems. J. Bacteriol. 92:502-509. 1966.-Leptospira pomona strain 3341 was grown in association with primary fetal bovine kidney (PBK) and human embryonic skin-muscle fibroblastic (HE) cells in Eagle's minimal essential medium (MEM) with 5% sheep serum. Growth curves of leptospires in PBK and HE cell cultures showed no substantial increase in growth above that obtained in Eagle's MEM in the absence of tissue culture cells. This suggested that no stimulatory growth factors for leptospires were produced by the tissue cells. Fibroblastic cells of the PBK monolayer showed separation, deterioration, and, finally, complete disintegration. Epithelial-like cells remained unaffected. HE cells showed the same cytopathic effect as PBK fibroblastic cells, indicating that this effect was not limited to PBK fibroblastic cells. Warthin-Starry stains of PBK and HE cell monolayers showed masses of leptospires adhering to fibroblastic cells, whereas only a few were seen on epithelial-like cells. Large numbers of leptospires on the surface of fibroblastic cells are very likely associated with the cytopathic effect. Dislodgment of leptospires from fibroblastic cells did not increase the total number of spirochetes in the culture. This indicated that leptospiral growth did not occur on the surface of these cells.     

Evaluation of the Sterifil Lysis-Filtration Blood Culture System

This paper describes the comparison of the Sterifil lysis-filtration (SLF) blood culture procedure with a standard Trypticase soy broth (TSB) technique. The lysing solutions employed in the SLF system, Triton X-100 (alkyl phenoxy polyethoxy ethanol) and sodium carbonate, were deleterious to most bacteria commonly encountered in bacteremia except staphylococci and enterococci. Candida was not adversely affected. There was a positive correlation between the tolerance of the microbial isolants to the lysing solutions and their recovery by the SLF technique. A total of 3,554 cultures were run in parallel and 398 isolants were obtained. Of 201 gram-positive isolants, 135 were recovered by both techniques, 43 were detected by the TSB technique only, and 23 were recovered only with the SLF method. In sharp contrast, of 168 gram-negative isolants, 28 were recovered in common, 130 were isolated only by TSB, and 10 were recovered only with the SLF method. The SLF method detected all cases of candidemia detected by the TSB method plus an additional 12 for a total of 29 cases. The SLF method, as currently described, is generally too toxic to bacteria for routine use in a clinical laboratory.     

Comparative Sensitivity of Various Cell Culture Systems for Isolation of Viruses from Wastewater and Fecal Samples

In efforts to define the most sensitive cell culture systems for recovery of viruses from wastewaters, 181 samples were inoculated in parallel into tube cultures of various cell types and were plaqued in bottle and petri dish cultures of three types of monkey kidney cells. Polioviruses were recovered most frequently in the RD line of human rhabdomyosarcoma cells, group A coxsackieviruses in RD and human fetal diploid kidney (HFDK) cells, group B coxsackieviruses in the BGM line of African green monkey kidney cells, echoviruses in RD and primary rhesus monkey kidney (RhMK) cells, and reoviruses in RhMK cells. BGM cells were unsatisfactory for recovery of viruses other than polioviruses and group B coxsackieviruses, and a line of fetal rhesus monkey kidney (MFK) was not a satisfactory substitute for primary RhMK. With RhMK cells, comparable numbers of virus isolations were made in tube cultures and in plaque assays conducted in bottle cultures, but with BGM and MFK cells, fewer isolations were made by plaquing than by inoculation of tube cultures. In comparative plaque assays on fecal samples under three different overlays in bottle and plate cultures of RhMK, BGM, and MFK cells, it was found that plaquing in the most sensitive system, RhMK, was less efficient for virus recovery than was inoculation of tube cultures of RhMK or HFDK cells. Overall, plaque assays performed in petri dishes in a CO₂ incubator yielded fewer virus isolates than did parallel plaque assays performed in closed bottle cultures. Other limitations of plaque assays for recovery of human enteric viruses are discussed.     

Epidemiological Evaluation of Blood Culture Patterns among Neonates Receiving Vancomycin

The objective of this study was to assess the frequency of blood culture (BC) collection among neonates who received vancomycin. Demographic, clinical, microbiologic, and pharmacy data were collected for 1275 neonates (postnatal age 0–27 days) who received vancomycin at an Intermountain Healthcare facility between 1/2006 and 9/2011. Neonates treated with vancomycin had a BC collected 94 % (n = 1198) of the time, of which 37 % (n = 448) grew one or more bacterial organisms (BC positive). Of these, 1 % (n = 5) grew methicillin-resistant Staphylococcus aureus (MRSA), 71 % (n = 320) grew coagulase-negative Staphylococci (CoNS), 9 % (n = 40) grew methicillin-sensitive Staphylococcus aureus (MSSA), and 22 % (n = 97) grew other bacterial species (total exceeds 100 % due to co-detection). In patients with negative BC or no BC, vancomycin therapy was extended beyond 72 h 52 % of the time. The median duration of vancomycin therapy for patients with a negative BC was 4 (IQR: 2–10) days. BCs were frequently obtained among neonates who received vancomycin. Vancomycin therapy beyond the conventional ‘empiric’ treatment window of 48–72 h was common without isolation of resistant gram-positive bacteria.     

热球菌目(Thermococcales)遗传操作系统研究与应用进展

热球菌目(Thermococcales)是一类分离自浅海热泉或者深海热液口的超嗜热微生物,包括火球菌(Pyrococcus)、热球菌(Thermococcus)、古老球菌(Palaeococcus)。研究其生命活动的分子机制,基因的功能等必须借助遗传操作系统。由于选择标记的限制,Thermococcales遗传操作系统落后于其他菌株。近年来,在Thermococcales发现了内源质粒并可以将其改造用作遗传工具。如在Thermococcus kodakarensis及Pyrococcus furious等菌株内都建立了成熟的遗传系统,并用于基因敲除以及基因表达。将就Thermococcales内源质粒的发现和遗传操作系统的发展与应用加以阐述。     

Aspergillosis of Biliary Tract After Liver Transplantation: A Case Report

We reported a case of aspergillosis presented as cholangitis in a patient after liver transplantation, even with prophylactic use of fluconazole. The patient had multiple predisposing factors, such as leukocytopenia, immunosuppressive drug therapy. He died 2 days after an exploratory laparotomy was carried out. The histopathologic finding of biopsy specimen from biliary tract was positive for Aspergillus. And the pathogen was identified as Aspergillus flavus by mycological culture and PCR. The patient was confirmed as a case of Aspergillosis of biliary tract that was responded poorly to fluconazole. This indicated that azole should have been switched to more effective antifungal agents at earlier stage when the patients responded poorly to the original treatment.     

Strong and High-Precision Manipulation of Nanoparticle with Graphene-Coated Fiber Systems

Two kinds of graphene-coated fiber systems are proposed and studied for optical trapping. Their plasmonic modes in uniform environment and close to the substrate are studied in the finite element method. The optical forces exerted on dielectric nanoparticle by these systems are calculated by standalone waveguide approximation. It is found that for the dielectric particle with diameter of 1 nm, the maximal optical forces generated by certain modes are more than 10⁷ fN/W whereas their force ranges are only one to several nanometers. These results may have important applications in strong and high-precision optical tweezers.

     

Evaluation of Antimicrobial Therapy of Blood Culture Positive Healthcare-Associated Infections in Children

Aim

Knowledge of the quality of antimicrobial therapy (AMT) used for invasive healthcare-associated infections (HAIs) in paediatrics is scarce. Influence of the final information about the isolated pathogen on the subsequent targeted AMT was investigated in our study.

Methods

Data on 149 children (0–17 years) with blood culture positive HAIs were collected. The causative microbes under investigation were Staphylococcus aureus, Staphylococcus epidermidis, streptococci, Gram negative rods, and mixed infections were likewise included. For adjusting the antimicrobial regimen, an expert panel evaluated the quality of the targeted AMT and the delay of 72 hours after final microbiology results. AMT was regarded as inappropriate if the pathogen was totally resistant to the used antimicrobials (i) or if the chosen therapy was of not optimal efficacy against the pathogen (ii).

Results

17% of the patients received inappropriate AMT. Half of these infections 13/26 (50%) were treated with an antimicrobial to which the isolate was resistant. Three (3/13, 23%) of these patients received antimicrobials which were totally ineffective according to in vitro data. Suboptimal or too broad spectrum AMT was administered to 13/26 (50%) patients. The most common causes of inappropriate use were the use of beta-lactams in oxacillin-resistant Staphylococcus epidermidis infections and vancomycin given in oxacillin-sensitive Staphylococcus aureus infections.

Conclusion

Approximately 17% of the selected cohort received inappropriate AMT. More attention should be paid to the appropriate use of antimicrobials, and training of prescribers should be urgently provided.     

Comparison of a Blood Culture System Containing Liquoid and Sucrose with Systems Containing Either Reagent Alone

By utilizing parallel culture methods on multiple blood specimens, three blood culture systems were evaluated for their ability to allow rapid recovery of organisms. The same base medium was used for all three systems. System or flask 1 contained base medium with Liquoid added, flask 2 contained base medium with sucrose added, and flask 3 contained both Liquoid and sucrose. During the study period, there were 259 patients considered to have positive blood cultures. Organism recovery was accomplished from flask 1 in 214 of these patients, from flask 2 in 180 cases, and from flask 3 in 257 instances. The use of anticoagulants and osmotic stabilizers in blood culture systems is discussed in detail.     

Evaluation of Three Cell Culture Systems as Substrates for Influenza Virus Assay

The propagation of various influenza virus strains in primary rhesus monkey kidney (RMK), primary hamster kidney (HK), and the MDCK-USD canine kidney (CK) cell cultures was compared. Virus-infected cultures were examined for cytopathic effect (CPE) and by performing the hemadsorption (HAd) technique. The highest HAd titers were found most often in HK, followed by RMK and CK. However, the CK cell line provided the best substrate for detecting CPE. Although influenza B strains tended to grow to higher titers, these strains produced less CPE than did the A strains.     

Superposition of Automatic and Voluntary Aspects of Grip Force Control in Humans during Object Manipulation

When moving grasped objects, people automatically modulate grip force (GF) with movement-dependent load force (LF) in order to prevent object slip. However, GF can also be modulated voluntarily as when squeezing an object. Here we investigated possible interactions between automatic and voluntary GF control. Participants were asked to generate horizontal cyclic movements (between 0.6 and 2.0 Hz) of a hand-held object that was restrained by an elastic band such that the load force (LF) reached a peak once per movement cycle, and to simultaneously squeeze the object at each movement reversal (i.e., twice per cycle). Participants also performed two control tasks in which they either only moved (between 0.6 and 2.0 Hz) or squeezed (between 1.2 and 4.0 Hz) the object. The extent to which GF modulation in the simultaneous task could be predicted from the two control tasks was assessed using power spectral analyses. At all frequencies, the GF power spectra from the simultaneous task exhibited two prominent components that occurred at the cycle frequency (ƒ) and at twice this frequency (2ƒ), whereas the spectra from the movement and squeeze control task exhibited only single peaks at ƒ and 2ƒ, respectively. At lower frequencies, the magnitudes of both frequency components in the simultaneous task were similar to the magnitudes of the corresponding components in the control tasks. However, as frequency increased, the magnitudes of both components in the simultaneous task were greater than the magnitudes of the corresponding control task components. Moreover, the phase relationship between the ƒ components of GF and LF began to drift from the value observed in the movement control task. Overall these results suggest that, at lower movement frequencies, voluntary and automatic GF control processes operate at different hierarchical levels. Several mechanisms are discussed to account for interaction effects observed at higher movement frequencies.     

A Mass Spectrometric Study for Comparative Analysis and Evaluation of Metabolite Recovery from Plasma by Various Solvent Systems

A solvent system that extracts a maximum number of metabolites belonging to diverse chemical classes from complex biofluids, such as plasma, may offer useful inputs to understand the metabolic and physiological state of an individual. The present study compared seven solvent systems for extraction of metabolites from plasma. The extracts were analyzed by mass spectrometry (MS) and MS/MS (MS2) using a quadrupole time-of-flight liquid chromatography/MS system in positive and negative modes of ionization. Metabolites with molecular mass below 400 were identified using Human Metabolome Database MS2 and MS search interfaces. The acetone/isopropanol (2:1) system yielded promising results in positive ionization mode, as the maximum number of MS and MS2 features was detected in the extract. It was found to be superior in extraction of various classes of metabolites, especially organic acids, nucleosides and nucleoside derivatives, and heterocyclic molecules. Glycerophosphocholines in the mass range of 400–700 were found to be efficiently extracted by the methanol/chloroform/water (8:1:1) system. In negative mode as well, the maximum number of MS2 features was detected in methanol/chloroform/water and acetone/isopropanol extracts. The fingerprints of molecular features obtained in the negative and positive modes differed from each other to a significant extent.