Conductometric Immunosensor for Escherichia coli O157:H7 Detection Based on Polyaniline/Zinc Oxide (PANI/ZnO) Nanocomposite

A conductometric immunosensor was developed for the detection of one of the most common foodborne pathogens, Escherichia coli O157:H7 (E. coli O157:H7), by conductometric sensing. The sensor was built based on a polyaniline/zinc oxide (PANI/ZnO) nanocomposite film spin-coated on a gold electrode. Then, it was modified with a monoclonal anti-E. coli O157:H7 antibody as a biorecognition element. The fabricated nanostructured sensor was able to quantify the pathogens under optimal detection conditions, within 30 min, and showed a good detection range from 101 to 104 CFU/mL for E. coli O157:H7 and a minimum detection limit of 4.8 CFU/mL in 0.1% peptone water. The sensor efficiency for detecting bacteria in food matrices was tested in ultra-heat-treated (UHT) skim milk. E. coli O157:H7 was detected at concentrations of 101 to 104 CFU/mL with a minimum detection limit of 13.9 CFU/mL. The novel sensor was simple, fast, highly sensitive with excellent specificity, and it had the potential for rapid sample processing. Moreover, this unique technique for bacterial detection could be applicable for food safety and quality control in the food sector as it offers highly reliable results and is able to quantify the target bacterium.

Specific Detection of Bovine Coronavirus N Protein with TaqMan Probe qRT-PCR

Background: Bovine Coronavirus (BCoV) can cause acute diarrhea in newborn calves and adult cattle. BCoV infection may cause losses to production by reduced weight gain, reduced milk yield. Several methods have been applied to detect and diagnose BCoV. However, each assay has its deficiency. Currently, real-time quantitative PCR (qRT-PCR) has been utilized to identify and quantify many viral pathogens since it is a highly sensitive. However, the technical assay varies due to normalization control of the signal with an internal standard, typically a housekeeping gene. The main objective of the present study to establish a novel TaqMan probe real-time PCR (qRT-PCR) for detecting BCoV.Materials, Methods & Results: The present study was aimed to establish a novel TaqMan probe real-time PCR (qRT-PCR) for detecting bovine coronaviruses (BCoV), and also to develop a diagnostic protocol which simplifies sample collection and processing. One pair of specific primers, one pair of universal primers and a TaqMan probe were designed from the known sequences of conserved nucleocapsid (N) protein of BCoV. Reaction systems of TaqMan qRT-PCR were optimized including concentrations of the primers and probe as well as annealing temperatures. Prior to optimizing the assay, the recombinant plasmids of pMD18-T-BCoV-N were successfully constructed to make standard curves. The sensitivity, specificity and reproducibility were evaluated on the TaqMan qRT-PCR, respectively. A total of 321 feces specimens collected from diarrheic calves were detected with this assay. The results showed the optimized reaction conditions for qRT-PCR were 14.5 μM/L primers, 19.5 μM/L probes and 45.0°C annealing temperatures. The established TaqMan qRT-PCR assay could specially detect BCoV without detecting any other viruses. Its minimum detection limit was 4.72 × 101 copies/μL. However, universal PCR could detect only 4.72 × 103 copies/μL. Its sensitivity was 100-fold stronger than universal PCR. In conclusion, this TaqMan qRT-PCR had excellent specificity, sensitivity and stability with a 100-fold sensitivity stronger than universal PCR. Minimum detection limit was 4.72 × 101 copies/μL. This method was a cost-effective method to diagnose diarrhea and distinguish pathogens in dairy farms.Discussion: In this study, the authors developed a quantitative real-time PCR (qRT-PCR) in this study based on the TaqMan probe of BCoV. This TaqMan qRT-PCR assay selected and used one pair of specific primers (BCoV-qF/BCoV-qR) and a specific TaqMan probe (BCoV-probe) targeting the conserved nucleocapsid (N) gene. The specificity of primers and probes was validated with Primer-BLAST. The specificity of the qRT-PCR was confirmed by the negative control and other six viruses. The findings demonstrated that TaqMan qRT-PCR could only detect BCoV. This verified the qRT-PCR had an excellent specificity. It is obvious that this TaqMan qRT-PCR assay can detect only BCoV with stronger sensitivity and reproducibility than other real-time PCR methods. The sensitivity test indicated the minimum detection limit of the TaqMan qRT-PCR was 4.72 × 101copies/μL, or 47.2 copies/μL. Sensitivity of the TaqMan qRT-PCR assay was increased by 100-fold as compared to universal PCR with a good inter-assay and intra-assay reproducibility. Thereby, based on the high sensitivity of the assay of this qRT-PCR assay it may be a cost-effective method to diagnose BCoV infections and indentify the etiologic agents of diarrhea syndrome in the dairy farms.

Mobile measurement of methane emissions from natural gas developments in northeastern British Columbia, Canada

North American leaders recently committed to reducing methane emissions from the oil and gas sector, but information on current emissions from upstream oil and gas developments in Canada are lacking. This study examined the occurrence of methane plumes in an area of unconventional natural gas development in northwestern Canada. In August to September 2015 we completed almost 8000 km of vehicle-based survey campaigns on public roads dissecting oil and gas infrastructure, such as well pads and processing facilities. We surveyed six routes 3–6 times each, which brought us past over 1600 unique well pads and facilities managed by more than 50 different operators. To attribute on-road plumes to oil- and gas-related sources we used gas signatures of residual excess concentrations (anomalies above background) less than 500 m downwind from potential oil and gas emission sources. All results represent emissions greater than our minimum detection limit of 0.59 g s−1 at our average detection distance (319 m). Unlike many other oil and gas developments in the US for which methane measurements have been reported recently, the methane concentrations we measured were close to normal atmospheric levels, except inside natural gas plumes. Roughly 47 % of active wells emitted methane-rich plumes above our minimum detection limit. Multiple sites that pre-date the recent unconventional natural gas development were found to be emitting, and we observed that the majority of these older wells were associated with emissions on all survey repeats. We also observed emissions from gas processing facilities that were highly repeatable. Emission patterns in this area were best explained by infrastructure age and type. Extrapolating our results across all oil and gas infrastructure in the Montney area, we estimate that the emission sources we located (emitting at a rate > 0.59 g s−1) contribute more than 111 800 t of methane annually to the atmosphere. This value exceeds reported bottom-up estimates of 78 000 t of methane for all oil and gas sector sources in British Columbia. Current bottom-up methods for estimating methane emissions do not normally calculate the fraction of emitting oil and gas infrastructure with thorough on-ground measurements. However, this study demonstrates that mobile surveys could provide a more accurate representation of the number of emission sources in an oil and gas development. This study presents the first mobile collection of methane emissions from oil and gas infrastructure in British Columbia, and these results can be used to inform policy development in an era of methane emission reduction efforts.

Mobile measurement of methane emissions from natural gas developments in Northeastern British Columbia, Canada

North American leaders recently committed to reducing methane emissions from the oil and gas sector, but information on current emissions from Canadian unconventional developments is lacking. This study examined the incidence of methane in an area of unconventional natural gas development in northwestern Canada. In August to September 2015 we completed almost 8000 km of vehicle-based survey campaigns on public roads dissecting developments that mainly access the Montney formation in northeastern British Columbia. Six survey routes were repeated 3–6 times and brought us past over 1600 unique well pads and facilities developed by more than 50 different operators. To attribute on-road plumes to infrastructural sources we used gas signatures of residual excess concentrations (anomalies above background) less than 500 m downwind from infrastructural sources. All results represent emissions greater than our minimum detection limit of 0.59 g/s at our average detection distance (319 m). Unlike many other developments in the US for which methane measurements have been reported recently, the methane concentrations we measured at surface were close to normal atmospheric levels, except inside natural gas plumes. Roughly 47 % of active wells emitted methane-rich plumes above our minimum detection limit. Abandoned and under-development well sites also emitted methane-rich plumes, but the incidence rate was below that of producing wells. Multiple sites that pre-date the recent unconventional Montney development were found to be emitting, and in general we observed that older infrastructure tended to emit more often (per unit) with comparable severity in terms of measured excess concentrations on-road. We also observed emissions from facilities of various types that were highly repeatable. Emission patterns in this area were best explained by infrastructure age and type. Extrapolating our results across the Montney development, we estimate that the emission sources we located (emitting at a rate > 0.59 g/s) contribute more than 111,800 tonnes of methane annually to the atmosphere. This value exceeds reported bottom-up estimates of 78,000 tonnes for all oil and gas sector sources in British Columbia, of which the Montney represents about 55 % of production. The results also demonstrate that mobile surveys could be used to exhaustively screen developments for super-emitters, because without our intensive 6-fold replication we could have used single-pass sampling to screen 80 % of Montney-related infrastructure. This is the first bottom-up study of fugitive emissions in the Canadian energy sector, and these results can be used to inform policy development in an era of methane emission reduction efforts.

A Study on Prevalence of Escherichia coli O157 with a Verified Method in Foods

The purpose of this study were to identify the presence of E.coli O157 and to determine its prevalence in foods which were collected from various restaurants, shops and markets in Istanbul. Also, validation of detection method of E. coli O157 in all food stuffs was carried out according to applicability, repeatability, and minimum detection limit (LOD) and false positive and negative analysis based on TS EN ISO 16654 standard method. The results showed that the prevalence of E. coli O157 in food was 2%, and its prevalence increased in April and May.

A pyrenesulfonyl-imidazolium derivative as a selective cyanide ion sensor in aqueous media

Unlocking of probe 2 with CN− ions leads to its minimum detection limit of 0.5 μM (13 ppb).

Low-power total reflection X-ray fluorescence spectrometer using diffractometer guide rail

An X-ray diffractometer (XRD) was modified to a low-power total reflection X-ray fluorescence (TXRF) spectrometer. This was realized by reducing the XRD tube power (3 kW) down to 10 W by a Spellman power supply. The present spectrometer consisted of a waveguide slit, Si-PIN detector, a goniometer and two Z-axis stages that were set on a diffractometer guide rail. This unit was easy in assembly. The first measurements with this spectrometer were presented. The minimum detection limit for Cr was estimated to be a few nanograms or at the level of 1013 atoms cm−2.