Accurate Estimation of Bicarbonate and Acetic Acid Concentrations with Wider Ranges in Anaerobic Media Using Classical FOS/TAC Titration Method

The determination of a volatile fatty acid content (FOS) and total alkalinity (TAC) can be carried out using Nordmann’s FOS/TAC titration method developed in the 1970s. This two-point titration (pH = 5 and 4.4) can be simply implemented and is widely employed by both the academic and industrial worlds. However, the present study proves that Nordmann’s method is only valid in limited ranges, since the titration of one FOS and TAC has an impact on the determination of the other, especially in extreme conditions. The present work develops a numerical tool with Scilab simulating the acid–base equilibria of titration. The program is efficient in predicting the experimental equivalent volumes obtained from Nordmann’s method with different combinations of sodium acetate and sodium bicarbonate contents. The mean absolute percentage errors (MAPE) between the simulation and experiment are below 7%. Two new formulas are developed, considering both equivalent volumes at pH = 5 and 4.4 to calibrate FOS and TAC values. The proposed formulas show their good performance in predicting various combinations of FOS and TAC contents in an anaerobic digestate at TAC ranging from 0 to 20,000 mg CaCO3·L−1 and FOS ranging from 0 to 31,000 mg HAc·L−1.

Characterization and Performance of Lactate-Feeding Consortia for Reductive Dechlorination of Trichloroethene

Understanding the underlying mechanism that drives the microbial community mediated by substrates is crucial to enhance the biostimulation in trichloroethene (TCE)-contaminated sites. Here, we investigated the performance of stable TCE-dechlorinating consortia by monitoring the variations in TCE-related metabolites and explored their underlying assembly mechanisms using 16S rDNA amplicon sequencing and bioinformatics analyses. The monitoring results indicated that three stable TCE-dechlorinating consortia were successfully enriched by lactate-containing anaerobic media. The statistical analysis results demonstrated that the microbial communities of the enrichment cultures changed along with time and were distinguished by their sample sources. The deterministic and stochastic processes were simultaneously responsible for shaping the TCE-dechlorinating community assembly. The indicator patterns shifted with the exhaustion of the carbon source and the pollutants, and the tceA-carrying Dehalococcoides, as an indicator for the final stage samples, responded positively to TCE removal during the incubation period. Pseudomonas, Desulforhabdus, Desulfovibrio and Methanofollis were identified as keystone populations in the TCE-dechlorinating process by co-occurrence network analysis. The results of this study indicate that lactate can be an effective substrate for stimulated bioremediation of TCE-contaminated sites, and the reduction of the stochastic forces or enhancement of the deterministic interventions may promote more effective biostimulation.

Processing of Positive Blood Cultures Using a Total Laboratory Automation System: Workflow and Clinical Validation

Automated systems for culture-based microbiology are in the early phase of implementation in clinical laboratories. Here, our objective was to evaluate the performance of the BD Kiestra Total Laboratory Automation (TLA) System for inoculation, incubation, and imaging of positive blood culture broth specimens. To optimize parameters for clinical testing, 56 clinical specimens were processed using both TLA and manual standard-of-care (SOC) methods. For TLA processing, 3 mL positive blood culture broth (35 VersaTREK: 19 aerobic, 16 anaerobic; 21 BD BACTEC: 15 aerobic, 6 anaerobic) was transferred to a no-additive vacutainer using a safety adapter and syringe. This aliquot was then placed on TLA for fully automated processing: 10 µL was inoculated to blood, chocolate, and MacConkey agar (Remel) and, for anaerobic bottles only, Brucella blood agar (Hardy Diagnostics). Kiestra cross-streak pattern 5 was optimal for obtaining isolated colonies and was superior to quadrant-streaking methods. Additional media types were added based on Gram stain results of the positive blood specimen: CandiSelect (Bio-Rad) and Sabouraud Dextrose (Remel) were added if yeast were identified, and Colistin Nalidixic Acid agar (Remel) for Gram stains with mixed Gram-positive and Gram-negative morphology. Plates were imaged at 6, 8, 10, 12, 18, 38, and 62 hours. Anaerobic media were placed by the system in a media stacker, incubated off-line, and then imaged on the TLA at 24, 48, and 72 hours. SOC cultures were evaluated after overnight incubation and on days 2 and 3. Organism identification was performed using MALDI-TOF MS (Bruker). Based on our evaluation, optimal parameters for clinical implementation of TLA were identified. Microbial growth was scant at 6 hours of incubation, but by 8 hours, small discrete colonies were observed for most aerobes. Thus, imaging parameters selected for routine clinical testing were 8, 18, and 38 hours for aerobic media and one image taken at 38 hours for anaerobic media. TLA and SOC culture results had 96% agreement (29 Gram positive, 12 Gram negative, 5 mixed, 6 yeast, 1 no growth). Two specimens with a second low-abundance bacterial population were observed on TLA that were not observed with SOC. Reproducibility of TLA was tested by processing 6 positive samples in triplicate and found to be 100%. There was no carryover or contamination noted between specimens processed simultaneously on TLA. To evaluate if implementation of TLA impacted epidemiology of positive blood cultures, we compared the 10 most common organisms recovered pre- and post-TLA implementation (August-November 2017 compared to August-November 2018). We found these organisms were not significantly impacted by TLA processing. In conclusion, automated processing of positive blood cultures improves laboratory workflows and facilitates faster workup at 8 hours of incubation. These results demonstrate that automation is a viable avenue for the processing of positive blood cultures.

Biodiversity of anaerobic cellulolytic bacteria in landfill sites

Landfills play an important role in the removal of waste from the surroundings. There is a limit to the types of waste that can be recycled and the landfill becomes the final method of waste disposal. Because waste constitutes a wide variety of materials, the microbial consortia that develop within a landfill will be equally varied, depending on the type of waste deposited, the temperature of the landfill and moisture content of the waste. The metabolism of these microbial consortia can result in products that are either harmful or beneficial. In order to increase the pool of knowledge on landfill microbiology, it is important to study the various consortia that inhabit the landfill to determine the various microbial interactions that occur and subsequently to manipulate these interactions to enhance the benefits of a landfill site and reduce the harmful effects. In this research, an attempt was made to isolate anaerobic cellulolytic bacteria from a landfill site. Six waste samples, varying in age were obtained over a period of two years. Samples were excavated from a maximum depth of 4m. Samples are processed in anaerobic, phosphate buffer and cultivated in various pre-reduced anaerobic media and incubated under anaerobic conditions. Samples were also collected from other potential anaerobic sites namely, anaerobic sludge, decomposing bagasse, compost, manure, rumen and pond sediment. Results of degradation of the cellulose source (Whatman No. 1 filter paper) indicated that it was possible to cultivate cellulose-degrading microorganisms from the landfill. Zones of clearing around colonies, which would be indicative of cellulose degradation on solid media, were not obtained. Samples from the anaerobic sludge, compost and rumen showed degradation of cellulose in liquid media but not on solid media. It is concluded that the solid media used was unsuitable for the cultivation of anaerobic, cellulolytic bacteria or that the anaerobic conditions employed were not adequate to initiate the growth of the anaerobic cellulolytic bacteria.

Bacteraemia during Direct Laryngoscopy and Endotracheal Intubation: A Study Using a Multiple Culture, Large Volume Technique

Bacteraemia secondary to orotracheal intubation has been reported to occur in 0-5.3% of patients. Bacteraemia detection is dependent upon several factors including the volume of blood per culture and the number of cultures. Prior studies used small volumes of blood and one or two cultures, and may therefore have underestimated the incidence of bacteraemia. Sixty-two adult patients who underwent direct laryngoscopy and endotracheal intubation were studied. Baseline blood cultures were sterile in all patients. After intubation, four blood cultures were obtained in ten minutes, with 10 ml being evenly divided between aerobic and anaerobic media. Two patients (3.2%) became bacteraemic. This is a lower incidence than occurs in association with other procedures for which The American Heart Association does not recommend administration of prophylactic antibiotics. Therefore, prophylactic antibiotics are not recommended prior to direct laryngoscopy. However, when a prophylactic antibiotic is administered prior to surgery, it would be best to administer the antibiotic prior to direct laryngoscopy and intubation.

Comparison of Six Commercially Available Transport Media for Maintenance of Serpulina (Treponema) Hyodysenteriae

Two anaerobic (A1 and A2), 1 selective (S1), and 3 conventional (C1, C2, and C3) transport media formulations were compared for their capacity to maintain the viability of Serpulina (Treponema) hyodysenteriae. Initial experiments compared the recovery of S. hyodysenteriae from pure cultures held in each transport medium for 0.5, 1, 2, 3, 5, and 7 days at −40 C, 4 C, 25 C, and 36 C. Subsequent experiments compared each transport medium for maintenance of S. hyodysenteriae in fecal specimens obtained from experimentally infected pigs after holding for up to 7 days at 25 C. In each experiment, the viability of S. hyodysenteriae in each transport medium incubated at each temperature and for each period was determined by inoculating the transport medium onto either trypticase soy agar with 5% sheep blood or selective BJ agar and incubating at 42 C anaerobically. Viability and fecal flora contamination were evaluated blindly after 2-, 4-, and 6-day incubation periods. At −40 C, recovery of viable S. hyodysenteriae from pure culture did not differ among the transport media from 0.5 to 7 days, and all of the transport media consistently maintained the viability of the spirochetes for 7 days. At 4 C, the anaerobic and selective transport media maintained the viability of pure cultures of S. hyodysenteriae significantly better than did the conventional transport media group at day 7 ( P = 0.019). At the same temperature, the anaerobic media maintained viability better than did the conventional media at 5 days ( P < 0.042). At 25 C, the anaerobic transport media were significantly better than the conventional transport media at maintaining the viability of pure cultures of the spirochetes at 2, 3, and 5 days ( P < 0.018) and were significantly better than the selective medium at 3 days (P = 0.012). At 36 C, the recovery of viable spirochetes was significantly better with the anaerobic transport media than with both the conventional media for days 2–7 ( P < 0.006) and the selective medium for days 3–7 ( P < 0.049). Fecal specimens held in transport media Al and C1, as a group, had significantly higher viability than those held in the other transport media formulations taken as a group at all incubation times, except 0.5 day ( P < 0.0046). Contamination of selective BJ medium by fecal flora was markedly higher after holding fecal specimens in conventional transport media than in anaerobic and selective transport media. In a dilution trial of a pure culture of spirochetes, transport media Al and A2 maintained the viability of 108 S. hyodysenteriae for 7 days; however, medium Al was 10–100-fold more effective than medium A2 when lower initial concentrations of spirochetes were sampled. In a dilution trial of a fecal specimen, medium Al maintained the viability of 101 spirochetes for 2.5 days compared with 103 with medium C1. Overall, media A1 and C1 were the most satisfactory transport media for recovery of viable S. hyodysenteriae from fecal specimens held at 25 C for up to 7 days.