Rapid detection of Escherichia coli O157:H7 using tunneling magnetoresistance biosensor

NASA Astrophysics Data System (ADS)

Wu, Yuanzhao; Liu, Yiwei; Zhan, Qingfeng; Liu, J. Ping; Li, Run-Wei

2017-05-01

A rapid method for the sensitive detection of bacteria using magnetic immunoassay, which are measured with a tunneling magnetoresistance (TMR) sensor, is described. For the measurement of Escherichia coli O157:H7 (E. coli O157:H7) bacteria, the target was labeled by magnetic beads through magnetic immunoassay. The magnetic beads produce a weak magnetic fringe field when external field is applied, thus induce the magnetoresistance change of TMR sensor. A detection limit of 100 CFU/mL E. coli O157:H7 bacteria in 5 hours was obtained. With its high sensitive and rapid detection scheme based on the TMR biosensor, the detection system is an excellent candidate suitable and promising for food safety and biomedical detection.

Volatile arsenic species released from Escherichia coli expressing the AsIII S-adenosylmethionine methyltransferase gene.

PubMed

Yuan, Chungang; Lu, Xiufen; Qin, Jie; Rosen, Barry P; Le, X Chris

2008-05-01

Biological systems, ranging from bacteria and fungi to humans, can methylate arsenic. Recent studies have suggested that the AsIII S-adenosylmethionine methyltransferase (arsM) gene in bacteria was responsible for the removal of arsenic as the volatile arsines from the bacteria. However, there has been no direct measure of the arsines released from bacteria cultures. We describe here an integrated system incorporating the bacterial incubation and volatile arsenic species analysis, and we demonstrate its application to the identification of the volatile arsines produced in bacterial cultures. The headspace of the bacterial cultures was purged with helium, and the volatile arsenic species were trapped in a chromatographic column immersed in liquid nitrogen. The cryogenically trapped arsines [AsH3, (CH3)AsH2, (CH3)2AsH, and (CH3)3As] were separated by gas chromatography and were detected by inductively coupled plasma mass spectrometry. A hydride generation system was coupled to the bacterial culture system, allowing for spiking standards and for generating calibration arsines necessary for quantitative analysis. Both bacteria containing the arsM gene or its variant arsMC2 gene were able to produce 400-500 ng of trimethylarsine. No trimethylarsine was detectable in bacteria lacking the arsM gene (containing the vector plasmid as negative control). These results confirm that arsM is responsible for releasing arsenic as volatile species from the arsenic-resistant bacteria. Our results also show traces of AsH3, CH3AsH2, and (CH3)2AsH in cultures of bacteria expressing arsM. The method detection limits for AsH3, CH3AsH2, (CH3)2AsH, and (CH3)3As were 0.5, 0.5, 0.7, and 0.6 pg, respectively. The ability to quantify trace levels of these volatile arsenic species makes it possible to study the biotransformation and biochemical roles of the evolution of these volatile arsenic species by biological systems.

Application of Artificial Neuro-Fuzzy Logic Inference System for Predicting the Microbiological Pollution in Fresh Water

NASA Astrophysics Data System (ADS)

Bouharati, S.; Benmahammed, K.; Harzallah, D.; El-Assaf, Y. M.

The classical methods for detecting the micro biological pollution in water are based on the detection of the coliform bacteria which indicators of contamination. But to check each water supply for these contaminants would be a time-consuming job and a qualify operators. In this study, we propose a novel intelligent system which provides a detection of microbiological pollution in fresh water. The proposed system is a hierarchical integration of an Artificial Neuro-Fuzzy Inference System (ANFIS). This method is based on the variations of the physical and chemical parameters occurred during bacteria growth. The instantaneous result obtained by the measurements of the variations of the physical and chemical parameters occurred during bacteria growth-temperature, pH, electrical potential and electrical conductivity of many varieties of water (surface water, well water, drinking water and used water) on the number Escherichia coli in water. The instantaneous result obtained by measurements of the inputs parameters of water from sensors.

A new microfluidic approach for the one-step capture, amplification and label-free quantification of bacteria from raw samples.

PubMed

Pereiro, Iago; Bendali, Amel; Tabnaoui, Sanae; Alexandre, Lucile; Srbova, Jana; Bilkova, Zuzana; Deegan, Shane; Joshi, Lokesh; Viovy, Jean-Louis; Malaquin, Laurent; Dupuy, Bruno; Descroix, Stéphanie

2017-02-01

A microfluidic method to specifically capture and detect infectious bacteria based on immunorecognition and proliferative power is presented. It involves a microscale fluidized bed in which magnetic and drag forces are balanced to retain antibody-functionalized superparamagnetic beads in a chamber during sample perfusion. Captured cells are then cultivated in situ by infusing nutritionally-rich medium. The system was validated by the direct one-step detection of Salmonella Typhimurium in undiluted unskimmed milk, without pre-treatment. The growth of bacteria induces an expansion of the fluidized bed, mainly due to the volume occupied by the newly formed bacteria. This expansion can be observed with the naked eye, providing simple low-cost detection of only a few bacteria and in a few hours. The time to expansion can also be measured with a low-cost camera, allowing quantitative detection down to 4 cfu (colony forming unit), with a dynamic range of 100 to 10 7 cfu ml -1 in 2 to 8 hours, depending on the initial concentration. This mode of operation is an equivalent of quantitative PCR, with which it shares a high dynamic range and outstanding sensitivity and specificity, operating at the live cell rather than DNA level. Specificity was demonstrated by controls performed in the presence of a 500× excess of non-pathogenic Lactococcus lactis . The system's versatility was demonstrated by its successful application to the detection and quantitation of Escherichia coli O157:H15 and Enterobacter cloacae . This new technology allows fast, low-cost, portable and automated bacteria detection for various applications in food, environment, security and clinics.

Remote-controlled robotic platform ORPHEUS as a new tool for detection of bacteria in the environment.

PubMed

Nejdl, Lukas; Kudr, Jiri; Cihalova, Kristyna; Chudobova, Dagmar; Zurek, Michal; Zalud, Ludek; Kopecny, Lukas; Burian, Frantisek; Ruttkay-Nedecky, Branislav; Krizkova, Sona; Konecna, Marie; Hynek, David; Kopel, Pavel; Prasek, Jan; Adam, Vojtech; Kizek, Rene

2014-08-01

Remote-controlled robotic systems are being used for analysis of various types of analytes in hostile environment including those called extraterrestrial. The aim of our study was to develop a remote-controlled robotic platform (ORPHEUS-HOPE) for bacterial detection. For the platform ORPHEUS-HOPE a 3D printed flow chip was designed and created with a culture chamber with volume 600 μL. The flow rate was optimized to 500 μL/min. The chip was tested primarily for detection of 1-naphthol by differential pulse voltammetry with detection limit (S/N = 3) as 20 nM. Further, the way how to capture bacteria was optimized. To capture bacterial cells (Staphylococcus aureus), maghemite nanoparticles (1 mg/mL) were prepared and modified with collagen, glucose, graphene, gold, hyaluronic acid, and graphene with gold or graphene with glucose (20 mg/mL). The most up to 50% of the bacteria were captured by graphene nanoparticles modified with glucose. The detection limit of the whole assay, which included capturing of bacteria and their detection under remote control operation, was estimated as 30 bacteria per μL. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Integrated optical biosensor for rapid detection of bacteria

NASA Astrophysics Data System (ADS)

Mathesz, Anna; Valkai, Sándor; Újvárosy, Attila; Aekbote, Badri; Sipos, Orsolya; Stercz, Balázs; Kocsis, Béla; Szabó, Dóra; Dér, András

2016-02-01

In medical diagnostics, rapid detection of pathogenic bacteria from body fluids is one of the basic issues. Most state-of-the-art methods require optical labeling, increasing the complexity, duration and cost of the analysis. Therefore, there is a strong need for developing selective sensory devices based on label-free techniques, in order to increase the speed, and reduce the cost of detection. In a recent paper, we have shown that an integrated optical Mach-Zehnder interferometer, a highly sensitive all-optical device made of a cheap photopolymer, can be used as a powerful lab-on-a-chip tool for specific, labelfree detection of proteins. By proper modifications of this technique, our interferometric biosensor was combined with a microfluidic system allowing the rapid and specific detection of bacteria from solutions, having the surface of the sensor functionalized by bacterium-specific antibodies. The experiments proved that the biosensor was able to detect Escherichia coli bacteria at concentrations of 106 cfu/ml within a few minutes, that makes our device an appropriate tool for fast, label-free detection of bacteria from body fluids such as urine or sputum. On the other hand, possible applications of the device may not be restricted to medical microbiology, since bacterial identification is an important task in microbial forensics, criminal investigations, bio-terrorism threats and in environmental studies, as well.

Integrated optical biosensor for rapid detection of bacteria

NASA Astrophysics Data System (ADS)

Mathesz, Anna; Valkai, Sándor; Újvárosy, Attila; Aekbote, Badri; Sipos, Orsolya; Stercz, Balázs; Kocsis, Béla; Szabó, Dóra; Dér, András

2015-12-01

In medical diagnostics, rapid detection of pathogenic bacteria from body fluids is one of the basic issues. Most state-of-the-art methods require optical labeling, increasing the complexity, duration and cost of the analysis. Therefore, there is a strong need for developing selective sensory devices based on label-free techniques, in order to increase the speed, and reduce the cost of detection. In a recent paper, we have shown that an integrated optical Mach-Zehnder interferometer, a highly sensitive all-optical device made of a cheap photopolymer, can be used as a powerful lab-on-a-chip tool for specific, labelfree detection of proteins. By proper modifications of this technique, our interferometric biosensor was combined with a microfluidic system allowing the rapid and specific detection of bacteria from solutions, having the surface of the sensor functionalized by bacterium-specific antibodies. The experiments proved that the biosensor was able to detect Escherichia coli bacteria at concentrations of 106 cfu/ml within a few minutes, that makes our device an appropriate tool for fast, label-free detection of bacteria from body fluids such as urine or sputum. On the other hand, possible applications of the device may not be restricted to medical microbiology, since bacterial identification is an important task in microbial forensics, criminal investigations, bio-terrorism threats and in environmental studies, as well.

Rapid detection of single bacteria in unprocessed blood using Integrated Comprehensive Droplet Digital Detection

PubMed Central

Kang, Dong-Ku; Ali, M. Monsur; Zhang, Kaixiang; Huang, Susan S.; Peterson, Ellena; Digman, Michelle A.; Gratton, Enrico; Zhao, Weian

2014-01-01

Blood stream infection or sepsis is a major health problem worldwide, with extremely high mortality, which is partly due to the inability to rapidly detect and identify bacteria in the early stages of infection. Here we present a new technology termed ‘Integrated Comprehensive Droplet Digital Detection’ (IC 3D) that can selectively detect bacteria directly from milliliters of diluted blood at single-cell sensitivity in a one-step, culture- and amplification-free process within 1.5–4 h. The IC 3D integrates real-time, DNAzyme-based sensors, droplet microencapsulation and a high-throughput 3D particle counter system. Using Escherichia coli as a target, we demonstrate that the IC 3D can provide absolute quantification of both stock and clinical isolates of E. coli in spiked blood within a broad range of extremely low concentration from 1 to 10,000 bacteria per ml with exceptional robustness and limit of detection in the single digit regime. PMID:25391809

Filamentous bacteria existence in aerobic granular reactors.

PubMed

Figueroa, M; Val del Río, A; Campos, J L; Méndez, R; Mosquera-Corral, A

2015-05-01

Filamentous bacteria are associated to biomass settling problems in wastewater treatment plants. In systems based on aerobic granular biomass they have been proposed to contribute to the initial biomass aggregation process. However, their development on mature aerobic granular systems has not been sufficiently studied. In the present research work, filamentous bacteria were studied for the first time after long-term operation (up to 300 days) of aerobic granular systems. Chloroflexi and Sphaerotilus natans have been observed in a reactor fed with synthetic wastewater. These filamentous bacteria could only come from the inoculated sludge. Thiothrix and Chloroflexi bacteria were observed in aerobic granular biomass treating wastewater from a fish canning industry. Meganema perideroedes was detected in a reactor treating wastewater from a plant processing marine products. As a conclusion, the source of filamentous bacteria in these mature aerobic granular systems fed with industrial effluents was the incoming wastewater.

Impact of Aerosol Dust on xMAP Multiplex Detection of Different Class Pathogens

PubMed Central

Kleymenov, Denis A.; Gushchin, Vladimir A.; Gintsburg, Alexander L.; Tkachuk, Artem P.

2017-01-01

Environmental or city-scale bioaerosol surveillance can provide additional value for biodefense and public health. Efficient bioaerosol monitoring should rely on multiplex systems capable of detecting a wide range of biologically hazardous components potentially present in air (bacteria, viruses, toxins and allergens). xMAP technology from LuminexTM allows multiplex bead-based detection of antigens or nucleic acids, but its use for simultaneous detection of different classes of pathogens (bacteria, virus, toxin) is questionable. Another problem is the detection of pathogens in complex matrices, e.g., in the presence of dust. In the this research, we developed the model xMAP multiplex test-system aiRDeTeX 1.0, which enables detection of influenza A virus, Adenovirus type 6 Salmonella typhimurium, and cholera toxin B subunit representing RNA virus, DNA virus, gram-negative bacteria and toxin respectively as model organisms of biologically hazardous components potentially present in or spreadable through the air. We have extensively studied the effect of matrix solution (PBS, distilled water), environmental dust and ultrasound treatment for monoplex and multiplex detection efficiency of individual targets. All targets were efficiently detectable in PBS and in the presence of dust. Ultrasound does not improve the detection except for bacterial LPS. PMID:29238328

Standoff detection: distinction of bacteria by hyperspectral laser induced fluorescence

NASA Astrophysics Data System (ADS)

Walter, Arne; Duschek, Frank; Fellner, Lea; Grünewald, Karin M.; Hausmann, Anita; Julich, Sandra; Pargmann, Carsten; Tomaso, Herbert; Handke, Jürgen

2016-05-01

Sensitive detection and rapid identification of hazardous bioorganic material with high sensitivity and specificity are essential topics for defense and security. A single method can hardly cover these requirements. While point sensors allow a highly specific identification, they only provide localized information and are comparatively slow. Laser based standoff systems allow almost real-time detection and classification of potentially hazardous material in a wide area and can provide information on how the aerosol may spread. The coupling of both methods may be a promising solution to optimize the acquisition and identification of hazardous substances. The capability of the outdoor LIF system at DLR Lampoldshausen test facility as an online classification tool has already been demonstrated. Here, we present promising data for further differentiation among bacteria. Bacteria species can express unique fluorescence spectra after excitation at 280 nm and 355 nm. Upon deactivation, the spectral features change depending on the deactivation method.

DETECTION OF AEROSOLIZED BACTERIA IN EXPIRED AIR SAMPLES FROM ASIAN ELEPHANTS (ELEPHAS MAXIMUS).

PubMed

Burke, Sophie M; Vogelnest, Larry; Thompson, Paul; Tovey, Euan R; Williamson, Peter

2017-06-01

Elephant-mediated transmission of tuberculosis is assumed to be similar to human models, which state close and prolonged contact with an infected individual is required for transmission. Although considered a risk factor for infection, several case studies have reported that close contact with an elephant is not always necessary for transmission, and the role of aerosolized bacteria remains unclear. To investigate aerosol-mediated transmission of pathogenic bacteria from elephants, a method for the detection of aerosols using an adapted sampling system was developed. A commensal bacterium was isolated from the upper respiratory tract of elephants ( Elephas maximus ) and was used as a proxy organism to detect aerosolized droplets in the sampling system. It was found that elephants are capable of producing aerosolized bacterial particles of a size small enough to remain airborne for prolonged periods and penetrate the lower regions of the human respiratory tract.

Volatile Arsenic Species Released from Escherichia coli Expressing the AsIII S-adenosylmethionine Methyltransferase Gene

PubMed Central

YUAN, CHUNGANG; LU, XIUFEN; QIN, JIE; ROSEN, BARRY P.; LE, X. CHRIS

2015-01-01

Biological systems, ranging from bacteria and fungi to humans, can methylate arsenic. Recent studies have suggested that the AsIII S-adenosylmethionine methyltransferase (arsM) gene in bacteria was responsible for the removal of arsenic as the volatile arsines from the bacteria. However, there has been no direct measure of the arsines released from bacteria cultures. We describe here an integrated system incorporating the bacterial incubation and volatile arsenic species analysis, and we demonstrate its application to the identification of the volatile arsines produced in bacterial cultures. The headspace of the bacterial cultures was purged with helium, and the volatile arsenic species were trapped in a chromatographic column immersed in liquid nitrogen. The cryogenically trapped arsines [AsH3, (CH3)AsH2, (CH3)2AsH, and (CH3)3As] were separated by gas chromatography and were detected by inductively coupled plasma mass spectrometry. A hydride generation system was coupled to the bacterial culture system, allowing for spiking standards and for generating calibration arsines necessary for quantitative analysis. Both bacteria containing the arsM gene or its variant arsMC2 gene were able to produce 400–500 ng of trimethylarsine. No trimethylarsine was detectable in bacteria lacking the arsM gene (containing the vector plasmid as negative control). These results confirm that arsM is responsible for releasing arsenic as volatile species from the arsenic-resistant bacteria. Our results also show traces of AsH3, CH3AsH2, and (CH3)2AsH in cultures of bacteria expressing arsM. The method detection limits for AsH3, CH3AsH2, (CH3)2AsH, and (CH3)3As were 0.5, 0.5, 0.7, and 0.6 pg, respectively. The ability to quantify trace levels of these volatile arsenic species makes it possible to study the biotransformation and biochemical roles of the evolution of these volatile arsenic species by biological systems. PMID:18522094

Comparative analysis of nitrifying bacteria associated with freshwater and marine aquaria.

PubMed Central

Hovanec, T A; DeLong, E F

1996-01-01

Three nucleic acid probes, two for autotrophic ammonia-oxidizing bacteria of the beta subdivision of the class Proteobacteria and one for alpha subdivision nitrite-oxidizing bacteria, were developed and used to study nitrifying bacterial phylotypes associated with various freshwater and seawater aquarium biofilters. Nitrosomonas europaea and related species were detected in all nitrifying seawater systems and accounted for as much as 20% of the total eubacterial rRNA. In contrast, nitrifying bacteria belonging to the beta-proteobacterial subdivision were detected in only two samples from freshwater aquaria showing vigorous nitrification rates. rRNA originating from nitrite-oxidizing alpha subdivision proteobacteria was not detected in samples from either aquarium environment. The data obtained indicate that chemolithotrophic ammonia oxidation in the freshwater aquaria was not due to beta-proteobacterial phylotypes related to members of the genus Nitrosomonas and their close relatives, the organisms usually implicated in freshwater nitrification. It is likely that nitrification in natural environments is even more complex than nitrification in these simple systems and is less well characterized with regard to the microorganisms responsible. PMID:8702281

Standoff detection and classification of bacteria by multispectral laser-induced fluorescence

NASA Astrophysics Data System (ADS)

Duschek, Frank; Fellner, Lea; Gebert, Florian; Grünewald, Karin; Köhntopp, Anja; Kraus, Marian; Mahnke, Peter; Pargmann, Carsten; Tomaso, Herbert; Walter, Arne

2017-04-01

Biological hazardous substances such as certain fungi and bacteria represent a high risk for the broad public if fallen into wrong hands. Incidents based on bio-agents are commonly considered to have unpredictable and complex consequences for first responders and people. The impact of such an event can be minimized by an early and fast detection of hazards. The presented approach is based on optical standoff detection applying laser-induced fluorescence (LIF) on bacteria. The LIF bio-detector has been designed for outdoor operation at standoff distances from 20 m up to more than 100 m. The detector acquires LIF spectral data for two different excitation wavelengths (280 and 355 nm) which can be used to classify suspicious samples. A correlation analysis and spectral classification by a decision tree is used to discriminate between the measured samples. In order to demonstrate the capabilities of the system, suspensions of the low-risk and non-pathogenic bacteria Bacillus thuringiensis, Bacillus atrophaeus, Bacillus subtilis, Brevibacillus brevis, Micrococcus luteus, Oligella urethralis, Paenibacillus polymyxa and Escherichia coli (K12) have been investigated with the system, resulting in a discrimination accuracy of about 90%.

Microbial community functional structure in response to antibiotics in pharmaceutical wastewater treatment systems.

PubMed

Zhang, Yu; Xie, Jianping; Liu, Miaomiao; Tian, Zhe; He, Zhili; van Nostrand, Joy D; Ren, Liren; Zhou, Jizhong; Yang, Min

2013-10-15

It is widely demonstrated that antibiotics in the environment affect microbial community structure. However, direct evidence regarding the impacts of antibiotics on microbial functional structures in wastewater treatment systems is limited. Herein, a high-throughput functional gene array (GeoChip 3.0) in combination with quantitative PCR and clone libraries were used to evaluate the microbial functional structures in two biological wastewater treatment systems, which treat antibiotic production wastewater mainly containing oxytetracycline. Despite the bacteriostatic effects of antibiotics, the GeoChip detected almost all key functional gene categories, including carbon cycling, nitrogen cycling, etc., suggesting that these microbial communities were functionally diverse. Totally 749 carbon-degrading genes belonging to 40 groups (24 from bacteria and 16 from fungi) were detected. The abundance of several fungal carbon-degrading genes (e.g., glyoxal oxidase (glx), lignin peroxidase or ligninase (lip), manganese peroxidase (mnp), endochitinase, exoglucanase_genes) was significantly correlated with antibiotic concentrations (Mantel test; P < 0.05), showing that the fungal functional genes have been enhanced by the presence of antibiotics. However, from the fact that the majority of carbon-degrading genes were derived from bacteria and diverse antibiotic resistance genes were detected in bacteria, it was assumed that many bacteria could survive in the environment by acquiring antibiotic resistance and may have maintained the position as a main player in nutrient removal. Variance partitioning analysis showed that antibiotics could explain 24.4% of variations in microbial functional structure of the treatment systems. This study provides insights into the impacts of antibiotics on microbial functional structure of a unique system receiving antibiotic production wastewater, and reveals the potential importance of the cooperation between fungi and bacteria with antibiotic resistance in maintaining the stability and performance of the systems. Copyright © 2013 Elsevier Ltd. All rights reserved.

PATHOGENICITY OF BIOFILM BACTERIA

EPA Science Inventory

There is a paucity of information concerning any link between the microorganisms commonly found in biofilms of drinking water systems and their impacts on human health. For bacteria, culture-based techniques detect only a limited number of the total microorganisms associated wit...

Multiresistant Bacteria Isolated from Chicken Meat in Austria

PubMed Central

Zarfel, Gernot; Galler, Herbert; Luxner, Josefa; Petternel, Christian; Reinthaler, Franz F.; Haas, Doris; Kittinger, Clemens; Grisold, Andrea J.; Pless, Peter; Feierl, Gebhard

2014-01-01

Multidrug resistant bacteria (MDR bacteria), such as extended spectrum beta-lactamase (ESBL) Enterobacteriaceae, methicillin resistant Staphylococcus aureus (MRSA), and vancomycin-resistant Enterococci (VRE), pose a challenge to the human health care system. In recent years, these MDR bacteria have been detected increasingly outside the hospital environment. Also the contamination of food with MDR bacteria, particularly of meat and meat products, is a concern. The aim of the study was to evaluate the occurrence of MDR bacteria in chicken meat on the Austrian market. For this study, 50 chicken meat samples were analysed. All samples originated from chickens slaughtered in Austrian slaughterhouses and were marked as produced in Austria. Samples were analysed for the presence of ESBL Enterobacteriaceae, methicillin resistant Staphylococci and VRE. Resistance genes of the isolated bacteria were characterised by PCR and sequencing. In the present study 26 ESBL producing E. coli, five mecA gene harbouring Staphylococci (but no MRSA), and four VRE were detected in chicken meat samples of Austrian origin. In 24 (48%) of the samples no ESBL Enterobacteriaceae, MRSA, methicillin resistant coagulase negative Staphylococcus (MRCNS) or VRE could be detected. None of the samples contained all three types of investigated multiresistant bacteria. In concordance to previous studies, CTX-M-1 and SHV-12 were the dominant ESBL genes. PMID:25485979

A new microfluidic approach for the one-step capture, amplification and label-free quantification of bacteria from raw samples† †Electronic supplementary information (ESI) available. See DOI: 10.1039/c6sc03880h Click here for additional data file. Click here for additional data file. Click here for additional data file. Click here for additional data file. Click here for additional data file. Click here for additional data file.

PubMed Central

Pereiro, Iago; Bendali, Amel; Tabnaoui, Sanae; Alexandre, Lucile; Srbova, Jana; Bilkova, Zuzana; Deegan, Shane; Joshi, Lokesh; Viovy, Jean-Louis; Malaquin, Laurent

2017-01-01

A microfluidic method to specifically capture and detect infectious bacteria based on immunorecognition and proliferative power is presented. It involves a microscale fluidized bed in which magnetic and drag forces are balanced to retain antibody-functionalized superparamagnetic beads in a chamber during sample perfusion. Captured cells are then cultivated in situ by infusing nutritionally-rich medium. The system was validated by the direct one-step detection of Salmonella Typhimurium in undiluted unskimmed milk, without pre-treatment. The growth of bacteria induces an expansion of the fluidized bed, mainly due to the volume occupied by the newly formed bacteria. This expansion can be observed with the naked eye, providing simple low-cost detection of only a few bacteria and in a few hours. The time to expansion can also be measured with a low-cost camera, allowing quantitative detection down to 4 cfu (colony forming unit), with a dynamic range of 100 to 107 cfu ml–1 in 2 to 8 hours, depending on the initial concentration. This mode of operation is an equivalent of quantitative PCR, with which it shares a high dynamic range and outstanding sensitivity and specificity, operating at the live cell rather than DNA level. Specificity was demonstrated by controls performed in the presence of a 500× excess of non-pathogenic Lactococcus lactis. The system's versatility was demonstrated by its successful application to the detection and quantitation of Escherichia coli O157:H15 and Enterobacter cloacae. This new technology allows fast, low-cost, portable and automated bacteria detection for various applications in food, environment, security and clinics. PMID:28626552

Combined Dielectrophoresis and Impedance Systems for Bacteria Analysis in Microfluidic On-Chip Platforms

PubMed Central

Páez-Avilés, Cristina; Juanola-Feliu, Esteve; Punter-Villagrasa, Jaime; del Moral Zamora, Beatriz; Homs-Corbera, Antoni; Colomer-Farrarons, Jordi; Miribel-Català, Pere Lluís; Samitier, Josep

2016-01-01

Bacteria concentration and detection is time-consuming in regular microbiology procedures aimed to facilitate the detection and analysis of these cells at very low concentrations. Traditional methods are effective but often require several days to complete. This scenario results in low bioanalytical and diagnostic methodologies with associated increased costs and complexity. In recent years, the exploitation of the intrinsic electrical properties of cells has emerged as an appealing alternative approach for concentrating and detecting bacteria. The combination of dielectrophoresis (DEP) and impedance analysis (IA) in microfluidic on-chip platforms could be key to develop rapid, accurate, portable, simple-to-use and cost-effective microfluidic devices with a promising impact in medicine, public health, agricultural, food control and environmental areas. The present document reviews recent DEP and IA combined approaches and the latest relevant improvements focusing on bacteria concentration and detection, including selectivity, sensitivity, detection time, and conductivity variation enhancements. Furthermore, this review analyses future trends and challenges which need to be addressed in order to successfully commercialize these platforms resulting in an adequate social return of public-funded investments. PMID:27649201

High-throughput living cell-based optical biosensor for detection of bacterial lipopolysaccharide (LPS) using a red fluorescent protein reporter system.

PubMed

Jiang, Hui; Jiang, Donglei; Shao, Jingdong; Sun, Xiulan; Wang, Jiasheng

2016-11-14

Due to the high toxicity of bacterial lipopolysaccharide (LPS), resulting in sepsis and septic shock, two major causes of death worldwide, significant effort is directed toward the development of specific trace-level LPS detection systems. Here, we report sensitive, user-friendly, high-throughput LPS detection in a 96-well microplate using a transcriptional biosensor system, based on 293/hTLR4A-MD2-CD14 cells that are transformed by a red fluorescent protein (mCherry) gene under the transcriptional control of an NF-κB response element. The recognition of LPS activates the biosensor cell, TLR4, and the co-receptor-induced NF-κB signaling pathway, which results in the expression of mCherry fluorescent protein. The novel cell-based biosensor detects LPS with specificity at low concentration. The cell-based biosensor was evaluated by testing LPS isolated from 14 bacteria. Of the tested bacteria, 13 isolated Enterobacteraceous LPSs with hexa-acylated structures were found to increase red fluorescence and one penta-acylated LPS from Pseudomonadaceae appeared less potent. The proposed biosensor has potential for use in the LPS detection in foodstuff and biological products, as well as bacteria identification, assisting the control of foodborne diseases.

Engineering Approaches for the Detection and Control of Orthopaedic Biofilm Infections

PubMed Central

Ehrlich, Garth D.; Stoodley, Paul; Kathju, Sandeep; Zhao, Yongjun; McLeod, Bruce R.; Balaban, Naomi; Hu, Fen Ze; Sotereanos, Nicholas G.; Costerton, J. William; Stewart, Philip S.; Post, J. Christopher; Lin, Qiao

2005-01-01

Artificial joints are subject to chronic infections associated with bacterial biofilms, which only can be eradicated by the traumatic removal of the implant followed by sustained intravenous antibiotic therapy. We have adopted an engineering approach to develop electrical–current-based approaches to bacterial eradication and microelectromechanical systems that could be embedded within the implanted joint to detect the presence of bacteria and to provide in situ treatment of the infection before a biofilm can form. In the former case we will examine the combined bactericidal effects of direct and indirect electrical fields in combination with antibiotic therapy. In the latter case, bacterial detection will occur by developing a microelectromechanical–systems-based biosensor that can “eavesdrop” on bacterial quorum–sensing-based communication systems. Treatment will be effected by the release of a cocktail of pharmaceutical reagents contained within integral reservoirs associated with the implant, including a molecular jamming signal that competitively binds to the bacteria’s quorum sensing receptors (which will “blind” the bacteria, preventing the production of toxins) and multiple high dose antibiotics to eradicate the planktonic bacteria. This approach is designed to take advantage of the relatively high susceptibility to antibiotics that planktonic bacteria display compared with biofilm envirovars. Here we report the development of a generic microelectromechanical systems biosensor that measures changes in internal viscosity in a base fluid triggered by a change in the external environment. PMID:16056027

Development of a rapid optic bacteria detecting system based on ATP bioluminescence

NASA Astrophysics Data System (ADS)

Liu, Jun Tao; Luo, JinPing; Liu, XiaoHong; Cai, XinXia

2014-12-01

A rapid optic bacteria detecting system based on the principle of Adenosine triphosphate(ATP) bioluminescence was presented in this paper. This system consisted of bioluminescence-based biosensor and the high-sensitivity optic meter. A photon counting photomultiplier tube (PMT) module was used to improve the detection sensitivity, and a NIOS II/f processor based on a Field Programmable Gate Array(FPGA) was used to control the system. In this work, Micrococcus luteus were chosen as the test sample. Several Micrococcus luteus suspension with different concentration was tested by both T2011 and plate counting method. By comparing the two group results, an calibration curve was obtained from the bioluminescence intensity for Micrococcus luteus in the range of 2.3×102 ~ 2.3×106 CFU/mL with a good correlation coefficient of 0.960. An impacting Air microorganism sampler was used to capture Airborne Bacteria, and 8 samples were collected in different place. The TBC results of 8 samples by T2011 were between 10 ~ 2×103 cfu/mL, consistent with that of plate counting method, which indicated that 8 samples were between 10 ~ 3×103 cfu/mL. For total airborne bacteria count was small, correlation coefficient was poor. Also no significant difference was found between T2011 and plate counting method by statistical analyses.

Vancomycin-resistant gene identification from live bacteria on an integrated microfluidic system by using low temperature lysis and loop-mediated isothermal amplification

PubMed Central

Chang, Wen-Hsin; Yu, Ju-ching; Yang, Sung-Yi; Lin, Yi-Cheng; Wang, Chih-Hung; You, Huey-Ling; Wu, Jiunn-Jong; Lee, Gwo-Bin

2017-01-01

Vancomycin-resistant Enterococcus (VRE) is a kind of enterococci, which shows resistance toward antibiotics. It may last for a long period of time and meanwhile transmit the vancomycin-resistant gene (vanA) to other bacteria. In the United States alone, the resistant rate of Enterococcus to vancomycin increased from a mere 0.3% to a whopping 40% in the past two decades. Therefore, timely diagnosis and control of VRE is of great need so that clinicians can prevent patients from becoming infected. Nowadays, VRE is diagnosed by antibiotic susceptibility test or molecular diagnosis assays such as matrix-assisted laser desorption ionization/time-of-flight mass spectrometry and polymerase chain reaction. However, the existing diagnostic methods have some drawbacks, for example, time-consumption, no genetic information, or high false-positive rate. This study reports an integrated microfluidic system, which can automatically identify the vancomycin resistant gene (vanA) from live bacteria in clinical samples. A new approach using ethidium monoazide, nucleic acid specific probes, low temperature chemical lysis, and loop-mediated isothermal amplification (LAMP) has been presented. The experimental results showed that the developed system can detect the vanA gene from live Enterococcus in joint fluid samples with detection limit as low as 10 colony formation units/reaction within 1 h. This is the first time that an integrated microfluidic system has been demonstrated to detect vanA gene from live bacteria by using the LAMP approach. With its high sensitivity and accuracy, the proposed system may be useful to monitor antibiotic resistance genes from live bacteria in clinical samples in the near future. PMID:28798845

Electrically biased GaAs/AlGaAs heterostructures for enhanced detection of bacteria

NASA Astrophysics Data System (ADS)

Aziziyan, Mohammad R.; Hassen, Walid M.; Dubowski, Jan J.

2016-03-01

We have examined the influence of electrical bias on immobilization of bacteria on the surface of GaAs/AlGaAs heterostructures, functionalized with an alkanethiol based architecture. A mixture of biotinylated polyethylene glycol (PEG) thiol and hexadecanethiol was applied to attach neutravidin and antibodies targeting specific immobilization of Legionella pneumophila. An electrochemical setup was designed to bias biofunctionalized samples with the potential measured versus silver/silver chloride reference electrode in a three electrode configuration system. The immobilization efficiency has been examined with fluorescence microscopy after tagging captured bacteria with fluorescein labeled antibodies. We demonstrate more than 2 times enhanced capture of Legionella pneumophila, suggesting the potential of electrically biased biochips to deliver enhanced sensitivity in detecting these bacteria.

Catalyzed Reporter Deposition-Fluorescence In Situ Hybridization Allows for Enrichment-Independent Detection of Microcolony-Forming Soil Bacteria

PubMed Central

Ferrari, Belinda C.; Tujula, Niina; Stoner, Kate; Kjelleberg, Staffan

2006-01-01

Advances in the growth of hitherto unculturable soil bacteria have emphasized the requirement for rapid bacterial identification methods. Due to the slow-growing strategy of microcolony-forming soil bacteria, successful fluorescence in situ hybridization (FISH) requires an rRNA enrichment step for visualization. In this study, catalyzed reporter deposition (CARD)-FISH was employed as an alternative method to rRNA enhancement and was found to be superior to conventional FISH for the detection of microcolonies that are cultivated by using the soil substrate membrane system. CARD-FISH enabled real-time identification of oligophilic microcolony-forming soil bacteria without the requirement for enrichment on complex media and the associated shifts in community composition. PMID:16391135

Current advances in molecular methods for detection of nitrite-dependent anaerobic methane oxidizing bacteria in natural environments.

PubMed

Chen, Jing; Dick, Richard; Lin, Jih-Gaw; Gu, Ji-Dong

2016-12-01

Nitrite-dependent anaerobic methane oxidation (n-damo) process uniquely links microbial nitrogen and carbon cycles. Research on n-damo bacteria progresses quickly with experimental evidences through enrichment cultures. Polymerase chain reaction (PCR)-based methods for detecting them in various natural ecosystems and engineered systems play a very important role in the discovery of their distribution, abundance, and biodiversity in the ecosystems. Important characteristics of n-damo enrichments were obtained and their key significance in microbial nitrogen and carbon cycles was investigated. The molecular methods currently used in detecting n-damo bacteria were comprehensively reviewed and discussed for their strengths and limitations in applications with a wide range of samples. The pmoA gene-based PCR primers for n-damo bacterial detection were evaluated and, in particular, several incorrectly stated PCR primer nucleotide sequences in the published papers were also pointed out to allow correct applications of the PCR primers in current and future investigations. Furthermore, this review also offers the future perspectives of n-damo bacteria based on current information and methods available for a better acquisition of new knowledge about this group of bacteria.

SERS substrates fabricated using ceramic filters for the detection of bacteria

NASA Astrophysics Data System (ADS)

Mosier-Boss, P. A.; Sorensen, K. C.; George, R. D.; Obraztsova, A.

2016-01-01

SERS substrates were fabricated by filtering either Ag or Au colloidal particles onto rigid, ceramic filters - onto which suspensions of bacteria were then filtered. SERS spectra of the bacteria were obtained using a Raman spectrometer that has an 'orbital raster scan' capability. It was shown that bacteria samples prepared in this manner were uniformly distributed onto the surface of the SERS substrate. The effect of common buffer systems on the SERS spectra was investigated and the utility of using the SERS technique for speciation of bacteria was explored.

COMPOSITE SAMPLING FOR DETECTION OF COLIFORM BACTERIA IN WATER SUPPLY

EPA Science Inventory

Low densities of coliform bacteria introduced into distribution systems may survive in protected habitats. These organisms may interfere with and cause confusion in the use of the coliforms as indicators of sewage contamination of drinking water. Methods of increasing the probabi...

Automatic bio-sample bacteria detection system

NASA Technical Reports Server (NTRS)

Chappelle, E. W.; Colburn, M.; Kelbaugh, B. N.; Picciolo, G. L.

1971-01-01

Electromechanical device analyzes urine specimens in 15 minutes and processes one sample per minute. Instrument utilizes bioluminescent reaction between luciferase-luciferin mixture and adenosine triphosphate (ATP) to determine number of bacteria present in the sample. Device has potential application to analysis of other body fluids.

Quorum-Sensing Signal-Response Systems in Gram-Negative Bacteria

PubMed Central

Papenfort, Kai; Bassler, Bonnie

2016-01-01

Abstract / Preface Bacteria use quorum sensing to orchestrate gene expression programmes that underlie collective behaviours. Quorum sensing relies on the production, release, detection and group-level response to extracellular signalling molecules, which are called autoinducers. Recent work has discovered new autoinducers in Gram-negative bacteria, shown how these molecules are recognized by cognate receptors, revealed new regulatory components that are embedded in canonical signalling circuits and identified novel regulatory network designs. In this Review we examine how, together, these features of quorum sensing signal–response systems combine to control collective behaviours in Gram-negative bacteria and we discuss the implications for host–microbial associations and antibacterial therapy. PMID:27510864

Semiochemicals released from five bacteria identified from animal wounds infested by primary screwworms and their effects on fly behavioral activity.

PubMed

Zhu, Junwei J; Chaudhury, Muhammad F; Durso, Lisa M; Sagel, Agustin; Skoda, Steven R; Jelvez-Serra, Nadia S; Santanab, Euzebio Goulart

2017-01-01

The Primary screwworm, Cochliomyia hominivorax (Coquerel), is a serious pest feeding on living flesh of any warm-blooded animal, including humans. It was eradicated from the United States in the early 1980s using the sterile male technique. However, it was recently detected in populations of wild deer and pets in the Florida Keys of the US. For monitoring purposes, screwworm flies are normally trapped using attractant bait with liver. However, there has been little effort to develop an efficient monitoring system for detection of screwworm flies using a specific synthetic attractant blend. Several studies have shown that odors from animal wound fluids attract screwworm adults, particularly gravid females. Bacteria associated with animal wounds have been identified that act as a major source for this attraction. To understand what volatiles attract screwworms we inoculated bovine blood with previously identified bacteria. We identified volatile chemicals released from the inoculated blood and other selected media over time and assessed the effect of those chemicals on behavioral activity of adult screwworm flies. A total of 7 volatile compounds were collected from bacteria incubated in either broth or blood using solid-phase microextraction, and their chemical structures were identified by their characteristic mass spectrum fragments and confirmed by retention times in comparison to those of synthetic standards via gas chromatograph combined mass spectrometry analyses. Five major volatiles including dimethyl disulfide, dimethyl trisulfide, phenol, p-cresol and indole were detected from a mixture of 5 bacteria incubated in blood. The ratios of volatiles released differed among different incubation media, time and individual bacteria. A synthetic mixture containing the five compounds was demonstrated to be attractive to adult screwworm flies both in laboratory assays and field trapping trials. The results obtained from this study may assist in developing an efficient trapping system using the identified attractant blend to detect the infestation of primary screwworms. This is also the first study to explore the complex systems in volatile release profiles from 5 bacteria isolated from screwworm-infested animal wounds that are incubated with different media and incubation time, as well as individual and multi-species bacterial communities.

Diagnostic Value of PCR Analysis of Bacteria and Fungi from Blood in Empiric-Therapy-Resistant Febrile Neutropenia ▿

PubMed Central

Nakamura, Akiko; Sugimoto, Yuka; Ohishi, Kohshi; Sugawara, Yumiko; Fujieda, Atsushi; Monma, Fumihiko; Suzuki, Kei; Masuya, Masahiro; Nakase, Kazunori; Matsushima, Yoshiko; Wada, Hideo; Katayama, Naoyuki; Nobori, Tsutomu

2010-01-01

This study aimed to assess the clinical utility of PCR for the analysis of bacteria and fungi from blood for the management of febrile neutropenic patients with hematologic malignancies. Using a PCR system able to detect a broad range of bacteria and fungi, we conducted a prospective pilot study of periodic analyses of blood from patients following intensive chemotherapy. When fever occurred, it was treated with empirical antibiotic therapy, basically without knowledge of the PCR results. In 23 febrile episodes during the neutropenic period, bacteria were detected by PCR in 11 cases, while the same species were identified by blood culture in 3 cases. In 10 out of 11 PCR-positive cases, fever could be managed by empirical therapy. In the empirical-therapy-resistant case, the identification of Stenotrophomonas maltophilia by PCR led to improvement of fever. No fungi were detected by PCR in febrile cases, while Aspergillus fumigatus was detected in one afebrile patient, several days before a clinical diagnosis was made. In subsequent sporadic PCR analyses in 15 cases of febrile neutropenia, bacteria were detected by both PCR and blood culture in 7 cases and by PCR alone in 6. Fungi were not detected. While fever was improved by empirical therapy in 12 out of the 13 PCR-positive cases, the identification of Pseudomonas aeruginosa by PCR in one therapy-resistant case contributed to the successful treatment of persistent fever. Our results indicate that PCR analysis of bacteria from blood provides essential information for managing empirical-therapy-resistant febrile neutropenia. PMID:20392911

[Significance of bacteria detection with filter paper method on diagnosis of diabetic foot wound infection].

PubMed

Zou, X H; Zhu, Y P; Ren, G Q; Li, G C; Zhang, J; Zou, L J; Feng, Z B; Li, B H

2017-02-20

Objective: To evaluate the significance of bacteria detection with filter paper method on diagnosis of diabetic foot wound infection. Methods: Eighteen patients with diabetic foot ulcer conforming to the study criteria were hospitalized in Liyuan Hospital Affiliated to Tongji Medical College of Huazhong University of Science and Technology from July 2014 to July 2015. Diabetic foot ulcer wounds were classified according to the University of Texas diabetic foot classification (hereinafter referred to as Texas grade) system, and general condition of patients with wounds in different Texas grade was compared. Exudate and tissue of wounds were obtained, and filter paper method and biopsy method were adopted to detect the bacteria of wounds of patients respectively. Filter paper method was regarded as the evaluation method, and biopsy method was regarded as the control method. The relevance, difference, and consistency of the detection results of two methods were tested. Sensitivity, specificity, positive predictive value, negative predictive value, and accuracy of filter paper method in bacteria detection were calculated. Receiver operating characteristic (ROC) curve was drawn based on the specificity and sensitivity of filter paper method in bacteria detection of 18 patients to predict the detection effect of the method. Data were processed with one-way analysis of variance and Fisher's exact test. In patients tested positive for bacteria by biopsy method, the correlation between bacteria number detected by biopsy method and that by filter paper method was analyzed with Pearson correlation analysis. Results: (1) There were no statistically significant differences among patients with wounds in Texas grade 1, 2, and 3 in age, duration of diabetes, duration of wound, wound area, ankle brachial index, glycosylated hemoglobin, fasting blood sugar, blood platelet count, erythrocyte sedimentation rate, C-reactive protein, aspartate aminotransferase, serum creatinine, and urea nitrogen (with F values from 0.029 to 2.916, P values above 0.05), while there were statistically significant differences among patients with wounds in Texas grade 1, 2, and 3 in white blood cell count and alanine aminotransferase (with F values 4.688 and 6.833 respectively, P <0.05 or P <0.01). (2) According to the results of biopsy method, 6 patients were tested negative for bacteria, and 12 patients were tested positive for bacteria, among which 10 patients were with bacterial number above 1×10(5)/g, and 2 patients with bacterial number below 1×10(5)/g. According to the results of filter paper method, 8 patients were tested negative for bacteria, and 10 patients were tested positive for bacteria, among which 7 patients were with bacterial number above 1×10(5)/g, and 3 patients with bacterial number below 1×10(5)/g. There were 7 patients tested positive for bacteria both by biopsy method and filter paper method, 8 patients tested negative for bacteria both by biopsy method and filter paper method, and 3 patients tested positive for bacteria by biopsy method but negative by filter paper method. Patients tested negative for bacteria by biopsy method did not tested positive for bacteria by filter paper method. There was directional association between the detection results of two methods ( P =0.004), i. e. if result of biopsy method was positive, result of filter paper method could also be positive. There was no obvious difference in the detection results of two methods ( P =0.250). The consistency between the detection results of two methods was ordinary (Kappa=0.68, P =0.002). (3) The sensitivity, specificity, positive predictive value, negative predictive value, and accuracy of filter paper method in bacteria detection were 70%, 100%, 1.00, 0.73, and 83.3%, respectively. Total area under ROC curve of bacteria detection by filter paper method in 18 patients was 0.919 (with 95% confidence interval 0-1.000, P =0.030). (4) There were 13 strains of bacteria detected by biopsy method, with 5 strains of Acinetobacter baumannii, 5 strains of Staphylococcus aureus, 1 strain of Pseudomonas aeruginosa, 1 strain of Streptococcus bovis, and 1 strain of bird Enterococcus . There were 11 strains of bacteria detected by filter paper method, with 5 strains of Acinetobacter baumannii, 3 strains of Staphylococcus aureus, 1 strain of Pseudomonas aeruginosa, 1 strain of Streptococcus bovis, and 1 strain of bird Enterococcus . Except for Staphylococcus aureus, the sensitivity and specificity of filter paper method in the detection of the other 4 bacteria were all 100%. The consistency between filter paper method and biopsy method in detecting Acinetobacter baumannii was good (Kappa=1.00, P <0.01), while that in detecting Staphylococcus aureus was ordinary (Kappa=0.68, P <0.05). (5) There was no obvious correlation between the bacteria number of wounds detected by filter paper method and that by biopsy method ( r =0.257, P =0.419). There was obvious correlation between the bacteria numbers detected by two methods in wounds with Texas grade 1 and 2 (with r values as 0.999, P values as 0.001). There was no obvious correlation between the bacteria numbers detected by two methods in wounds with Texas grade 3 ( r =-0.053, P =0.947). Conclusions: The detection result of filter paper method is in accordance with that of biopsy method in the determination of bacterial infection, and it is of great importance in the diagnosis of local infection of diabetic foot wound.

Rapid antibiotic efficacy screening with aluminum oxide nanoporous membrane filter-chip and optical detection system.

PubMed

Tsou, Pei-Hsiang; Sreenivasappa, Harini; Hong, Sungmin; Yasuike, Masayuki; Miyamoto, Hiroshi; Nakano, Keiyo; Misawa, Takeyuki; Kameoka, Jun

2010-09-15

We have developed a filter-chip and optical detection system for rapid antibiotic efficacy screening. The filter-chip consisted of a 1-mL reservoir and an anodic aluminum oxide (AAO) nanoporous membrane. Sample solution with liquid growth media, bacteria, and antibiotics was incubated in the reservoir for a specific period of time. The number of live bacteria on the surface of membrane was counted after the incubation with antibiotics and filtration. Using this biosensing system, we have demonstrated a 1-h antibiotic screening for patients' clinical samples, significantly faster than the conventional antibiotic susceptibility tests that typically take more than 24h. This rapid screening nature makes the filter-chip and detection system ideal for tailoring antibiotic treatment to individual patients by reducing the microbial antibiotic resistance, and improving the survival rate for patients suffering from postoperative infections. Published by Elsevier B.V.

Decrease in zinc adsorption onto soil in the presence of EPS-rich and EPS-poor Pseudomonas aureofaciens.

PubMed

Drozdova, O Yu; Pokrovsky, O S; Lapitskiy, S A; Shirokova, L S; González, A G; Demin, V V

2014-12-01

The adsorption of Zn onto the humic and illuvial horizons of the podzol soil in the presence of soil bacteria was studied using a batch-reactor technique as a function of the pH (from 2 to 9) and the Zn concentration in solution (from 0.076mM to 0.760mM). Exopolysaccharides-forming aerobic heterotrophs Pseudomonas aureofaciens were added at 0.1 and 1.0gwetL(-1) concentrations to two different soil horizons, and Zn adsorption was monitored as a function of the pH and the dissolved-Zn concentration. The pH-dependent adsorption edge demonstrated more efficient Zn adsorption by the humic horizon than the mineral horizon at otherwise similar soil concentrations. The Zn adsorption onto the EPS-poor strain was on slightly lower than that onto EPS-rich bacteria. Similar differences in the adsorption capacities between the soil and bacteria were also detected by "langmuirian" constant-pH experiments conducted in soil-Zn and bacteria-Zn binary systems. The addition of 0.1gwetL(-1)P. aureofaciens to a soil-bacteria system (4gdryL(-1)soil) resulted in statistically significant decrease in the adsorption yield, which was detectable from both the pH-dependent adsorption edge and the constant-pH isotherm experiments. Increasing the amount of added bacteria to 1gwetL(-1) further decreased the overall adsorption in the full range of the pH. This decrease was maximal for the EPS-rich bacteria and minimal for the EPS-poor bacteria (a factor of 2.8 and 2.2 at pH=6.9, respectively). These observations in binary and ternary systems were further rationalized by linear-programming modeling of surface equilibria that revealed the systematic differences in the number of binding sites and the surface-adsorption constant of zinc onto the two soil horizons with and without bacteria. The main finding of this work is that the adsorption of Zn onto the humic soil-bacteria system is lower than that in pure, bacteria-free soil systems. This difference is statistically significant (p<0.05). As such, EPS-rich bacteria are capable of efficiently shielding the soil particles from heavy-metal adsorption. The removal efficiency of heavy metals in an abiotic organic-rich soil system should therefore be significantly higher than that in the presence of bacteria. This effect can be explained by the shielding of strongly bound metal sites on the organic-rich soil particles by inert bacterial exopolysaccharides. Copyright © 2014 Elsevier Inc. All rights reserved.

Cultivation, detection, and ecophysiology of anaerobic ammonium-oxidizing bacteria.

PubMed

Kartal, Boran; Geerts, Wim; Jetten, Mike S M

2011-01-01

Anaerobic ammonium-oxidizing (anammox) bacteria oxidize ammonium with nitrite under anoxic conditions. The anammox process is currently used to remove ammonium from wastewater and contributes significantly to the loss of fixed nitrogen from the oceans. In this chapter, we focus on the ecophysiology of anammox bacteria and describe new methodologies to grow these microorganisms. Now, it is possible to enrich anammox bacteria up to 95% with a membrane bioreactor that removes forces of selection for fast settling aggregates and facilitates the growth of planktonic cells. The biomass from this system has a high anaerobic ammonium oxidation rate (50 fmol NH(4)(+) · cell(-1) day(-1)) and is suitable for many ecophysiological and molecular experiments. A high throughput Percoll density gradient centrifugation protocol may be applied on this biomass for further enrichment (>99.5%) of anammox bacteria. Furthermore, we provide an up-to-date list of commonly used primers and introduce protocols for quantification and detection of functional genes of anammox bacteria in their natural environment. Copyright © 2011 Elsevier Inc. All rights reserved.

An investigation of the well-water quality: immunosensor for pathogenic Pseudomonas aeruginosa detection based on antibody-modified poly(pyrrole-3 carboxylic acid) screen-printed carbon electrode.

PubMed

Bekir, Karima; Bousimma, Feriel; Barhoumi, Houcine; Fedhila, Kais; Maaref, Abderrazak; Bakhrouf, Amina; Ben Ouada, Hafedh; Namour, Philippe; Jaffrezic-Renault, Nicole; Ben Mansour, Hedi

2015-12-01

In this report, we describe a new immunosensor designed for the detection and the quantification of Pseudomonas aeruginosa bacteria in water. The developed biosensing system was based on the immobilization of purified polyclonal anti P. aeruginosa antibodies on electropolymerized poly(pyrrole-3-carboxylic acid)/glassy carbon electrode. The building of the immunosensor step by step was evaluated by electrochemical measurements such as cyclic voltammetry (CV) and impedance spectroscopy (EIS). The electrochemical signature of the immunosensor was established by the change of the charge transfer resistance when the bacteria suspended in solution became attached to the immobilized antibodies. As a result, stable and high sensitive impedimetric immunosensor was obtained with a sensitivity of 0.19 kΩ/decade defined in the linear range from 10(1) to 10(7) CFU/mL of cellular concentrations. A low detection limit was obtained for the P. aeruginosa bacteria and a high selectivity when other bacteria were occasioned as well as Escherichia coli. The developed immunosensor was applied in detecting pathogenic P. aeruginosa in well-water.

Peptidoglycan recognition proteins in Drosophila immunity.

PubMed

Kurata, Shoichiro

2014-01-01

Innate immunity is the front line of self-defense against infectious non-self in vertebrates and invertebrates. The innate immune system is mediated by germ-line encoding pattern recognition molecules (pathogen sensors) that recognize conserved molecular patterns present in the pathogens but absent in the host. Peptidoglycans (PGN) are essential cell wall components of almost all bacteria, except mycoplasma lacking a cell wall, which provides the host immune system an advantage for detecting invading bacteria. Several families of pattern recognition molecules that detect PGN and PGN-derived compounds have been indentified, and the role of PGRP family members in host defense is relatively well-characterized in Drosophila. This review focuses on the role of PGRP family members in the recognition of invading bacteria and the activation and modulation of immune responses in Drosophila. Copyright © 2013 Elsevier Ltd. All rights reserved.

Regrowth of potential opportunistic pathogens and algae in reclaimed-water distribution systems.

PubMed

Jjemba, Patrick K; Weinrich, Lauren A; Cheng, Wei; Giraldo, Eugenio; Lechevallier, Mark W

2010-07-01

A study of the quality of reclaimed water in treated effluent, after storage, and at three points in the distribution system of four plants in California, Florida, Massachusetts, and New York was conducted for 1 year. The plants had different treatment processes (conventional versus membrane bioreactor), production capacities, and methods for storage of the water, and the intended end uses of the water were different. The analysis focused on the occurrence of indicator bacteria (heterotrophic bacteria, coliforms, Escherichia coli, and enterococci) and opportunistic pathogens (Aeromonas spp., enteropathogenic E. coli O157:H7, Legionella spp., Mycobacterium spp., and Pseudomonas spp.), as well as algae. Using immunological methods, E. coli O157:H7 was detected in the effluent of only one system, but it was not detected at the sampling points, suggesting that its survival in the system was poor. Although all of the treatment systems effectively reduced the levels of bacteria in the effluent, bacteria regrew in the reservoir and distribution systems because of the loss of residual disinfectant and high assimilable organic carbon levels. In the systems with open reservoirs, algal growth reduced the water quality by increasing the turbidity and accumulating at the end of the distribution system. Opportunistic pathogens, notably Aeromonas, Legionella, Mycobacterium, and Pseudomonas, occurred more frequently than indicator bacteria (enterococci, coliforms, and E. coli). The Mycobacterium spp. were very diverse and occurred most frequently in membrane bioreactor systems, and Mycobacterium cookii was identified more often than the other species. The public health risk associated with these opportunistic pathogens in reclaimed water is unknown. Collectively, our results show the need to develop best management practices for reclaimed water to control bacterial regrowth and degradation of water before it is utilized at the point of use.

Regrowth of Potential Opportunistic Pathogens and Algae in Reclaimed-Water Distribution Systems ▿

PubMed Central

Jjemba, Patrick K.; Weinrich, Lauren A.; Cheng, Wei; Giraldo, Eugenio; LeChevallier, Mark W.

2010-01-01

A study of the quality of reclaimed water in treated effluent, after storage, and at three points in the distribution system of four plants in California, Florida, Massachusetts, and New York was conducted for 1 year. The plants had different treatment processes (conventional versus membrane bioreactor), production capacities, and methods for storage of the water, and the intended end uses of the water were different. The analysis focused on the occurrence of indicator bacteria (heterotrophic bacteria, coliforms, Escherichia coli, and enterococci) and opportunistic pathogens (Aeromonas spp., enteropathogenic E. coli O157:H7, Legionella spp., Mycobacterium spp., and Pseudomonas spp.), as well as algae. Using immunological methods, E. coli O157:H7 was detected in the effluent of only one system, but it was not detected at the sampling points, suggesting that its survival in the system was poor. Although all of the treatment systems effectively reduced the levels of bacteria in the effluent, bacteria regrew in the reservoir and distribution systems because of the loss of residual disinfectant and high assimilable organic carbon levels. In the systems with open reservoirs, algal growth reduced the water quality by increasing the turbidity and accumulating at the end of the distribution system. Opportunistic pathogens, notably Aeromonas, Legionella, Mycobacterium, and Pseudomonas, occurred more frequently than indicator bacteria (enterococci, coliforms, and E. coli). The Mycobacterium spp. were very diverse and occurred most frequently in membrane bioreactor systems, and Mycobacterium cookii was identified more often than the other species. The public health risk associated with these opportunistic pathogens in reclaimed water is unknown. Collectively, our results show the need to develop best management practices for reclaimed water to control bacterial regrowth and degradation of water before it is utilized at the point of use. PMID:20453149

SERS substrates fabricated using ceramic filters for the detection of bacteria.

PubMed

Mosier-Boss, P A; Sorensen, K C; George, R D; Obraztsova, A

2016-01-15

SERS substrates were fabricated by filtering either Ag or Au colloidal particles onto rigid, ceramic filters - onto which suspensions of bacteria were then filtered. SERS spectra of the bacteria were obtained using a Raman spectrometer that has an 'orbital raster scan' capability. It was shown that bacteria samples prepared in this manner were uniformly distributed onto the surface of the SERS substrate. The effect of common buffer systems on the SERS spectra was investigated and the utility of using the SERS technique for speciation of bacteria was explored. Published by Elsevier B.V.

[The hazards of hospitals and selected public buildings of Legionella pneumophila].

PubMed

Sikora, Agnieszka; Kozioł-Montewka, Maria; Wójtowicz-Bobin, Małgorzata; Gładysz, Iwona; Dobosz, Paulina

2013-11-01

The registered infection and outbreaks of epidemic tend to monitor potential reservoirs of Legionella infection. According to the Act of 29 March 2007 on the requirements for the quality of water intended for human consumption are required to test for the presence and number of Legionella in the water system of hospitals. In case of detection of L. pneumophila serogroup 1 (SG 1) or increased above normal number other serogroups of bacteria it is necessary to eradicate these bacteria from the water system. The aim of this study was to assess the degree of contamination of the water supply system of selected public buildings and analyze the effectiveness of disinfection methods for the elimination of L. pneumophila in hot water systems. The materials for this study were hot and cold water samples which were collected from the water supply system of 23 different objects. Enumeration of Legionella bacteria in water samples was determined by membrane filtration (FM) and/or by surface inoculation methods according to the standards: PN-ISO 11731: 2002: "The quality of the water. Detection and enumeration of Legionella" and PN-EN ISO 11731-2: 2008: "Water quality--Detection and enumeration of Legionella--Part 2: Methodology of membrane filtration for water with a small number of bacteria". L. pneumophila was present in 164 samples of hot water, which accounted for 76.99%. In all tested water samples L. pneumophila SG 2-14 strains were detected. The most virulent strain--L. pneumophila SG 1 was not detected. In examined 23 objects in 12 of L. pneumophila exceed acceptable levels > 100 CFU/100 ml. The presence of L. pneumophila SG 2-14 demonstrated in all examined objects, indicating the risk of infection, and the need for permanent monitoring of the water system supply. The thermal disinfection is the most common, inexpensive, and effective method of control of L. pneumophila used in examined objects, but does not eliminate bacterial biofilm. Disinfection using the filters stopped of L. pneumophila, and was the method of complementary thermal disinfection. Chlorine dioxide is a very effective biocide for large numbers of L. pneumophila in water systems.

Modeling and analysis of a microresonating biosensor for detection of Salmonella bacteria in human blood.

PubMed

Bahadoran, Mahdi; Noorden, Ahmad Fakhrurrazi Ahmad; Chaudhary, Kashif; Mohajer, Faeze Sadat; Aziz, Muhammad Safwan; Hashim, Shahrin; Ali, Jalil; Yupapin, Preecha

2014-07-18

A new photonics biosensor configuration comprising a Double-side Ring Add-drop Filter microring resonator (DR-ADF) made from SiO2-TiO2 material is proposed for the detection of Salmonella bacteria (SB) in blood. The scattering matrix method using inductive calculation is used to determine the output signal's intensities in the blood with and without presence of Salmonella. The change in refractive index due to the reaction of Salmonella bacteria with its applied antibody on the flagellin layer loaded on the sensing and detecting microresonator causes the increase in through and dropper port's intensities of the output signal which leads to the detection of SB in blood. A shift in the output signal wavelength is observed with resolution of 0.01 nm. The change in intensity and shift in wavelength is analyzed with respect to the change in the refractive index which contributes toward achieving an ultra-high sensitivity of 95,500 nm/RIU which is almost two orders higher than that of reported from single ring sensors and the limit of detection is in the order of 1 × 10(-8) RIU. In applications, such a system can be employed for a high sensitive and fast detection of bacteria.

Modeling and Analysis of a Microresonating Biosensor for Detection of Salmonella Bacteria in Human Blood

PubMed Central

Bahadoran, Mahdi; Noorden, Ahmad Fakhrurrazi Ahmad; Chaudhary, Kashif; Mohajer, Faeze Sadat; Aziz, Muhammad Safwan; Hashim, Shahrin; Ali, Jalil; Yupapin, Preecha

2014-01-01

A new photonics biosensor configuration comprising a Double-side Ring Add-drop Filter microring resonator (DR-ADF) made from SiO2-TiO2 material is proposed for the detection of Salmonella bacteria (SB) in blood. The scattering matrix method using inductive calculation is used to determine the output signal's intensities in the blood with and without presence of Salmonella. The change in refractive index due to the reaction of Salmonella bacteria with its applied antibody on the flagellin layer loaded on the sensing and detecting microresonator causes the increase in through and dropper port's intensities of the output signal which leads to the detection of SB in blood. A shift in the output signal wavelength is observed with resolution of 0.01 nm. The change in intensity and shift in wavelength is analyzed with respect to the change in the refractive index which contributes toward achieving an ultra-high sensitivity of 95,500 nm/RIU which is almost two orders higher than that of reported from single ring sensors and the limit of detection is in the order of 1 × 10−8 RIU. In applications, such a system can be employed for a high sensitive and fast detection of bacteria. PMID:25046015

Biosensor for remote monitoring of airborne toxins

NASA Astrophysics Data System (ADS)

Knopf, George K.; Bassi, Amarjeet S.; Singh, Shikha; Macleod, Roslyn

1999-12-01

The rapid detection of toxic contaminants released into the air by chemical processing facilities is a high priority for many manufacturers. This paper describes a novel biosensor for the remote monitoring of toxic sites. The proposed biosensor is a measurement system that employs immobilized luminescent Vibrio fisheri bacteria to detect airborne contaminants. The presence of toxic chemicals will lead to a detectable decrease in the intensity of light produced by the bacteria. Both cellular and environmental factors control the bioluminescence of these bacteria. Important design factors are the appropriate cell growth media, environmental toxicity, oxygen and cell concentrations. The luminescent bacteria are immobilized on polyvinyl alcohol (PVA) gels and placed inside a specially constructed, miniature flow cell which houses a transducer, power source, and transmitter to convert the light signal information into radio frequencies that are picked up by a receiver at a remote location. The biosensor prototype is designed to function either as a single unit mounted on an exploratory robot or numerous units spatially distributed throughout a contaminated environment for remote sensing applications.

A novel, multiplex, real-time PCR-based approach for the detection of the commonly occurring pathogenic fungi and bacteria.

PubMed

Horváth, Ádám; Pető, Zoltán; Urbán, Edit; Vágvölgyi, Csaba; Somogyvári, Ferenc

2013-12-23

Polymerase chain reaction (PCR)-based techniques are widely used to identify fungal and bacterial infections. There have been numerous reports of different, new, real-time PCR-based pathogen identification methods although the clinical practicability of such techniques is not yet fully clarified.The present study focuses on a novel, multiplex, real-time PCR-based pathogen identification system developed for rapid differentiation of the commonly occurring bacterial and fungal causative pathogens of bloodstream infections. A multiplex, real-time PCR approach is introduced for the detection and differentiation of fungi, Gram-positive (G+) and Gram-negative (G-) bacteria. The Gram classification is performed with the specific fluorescence resonance energy transfer (FRET) probes recommended for LightCycler capillary real-time PCR. The novelty of our system is the use of a non-specific SYBR Green dye instead of labelled anchor probes or primers, to excite the acceptor dyes on the FRET probes. In conjunction with this, the use of an intercalating dye allows the detection of fungal amplicons.With the novel pathogen detection system, fungi, G + and G- bacteria in the same reaction tube can be differentiated within an hour after the DNA preparation via the melting temperatures of the amplicons and probes in the same tube. This modified FRET technique is specific and more rapid than the gold-standard culture-based methods. The fact that fungi, G + and G- bacteria were successfully identified in the same tube within an hour after the DNA preparation permits rapid and early evidence-based management of bloodstream infections in clinical practice.

A new analytical platform based on field-flow fractionation and olfactory sensor to improve the detection of viable and non-viable bacteria in food.

PubMed

Roda, Barbara; Mirasoli, Mara; Zattoni, Andrea; Casale, Monica; Oliveri, Paolo; Bigi, Alessandro; Reschiglian, Pierluigi; Simoni, Patrizia; Roda, Aldo

2016-10-01

An integrated sensing system is presented for the first time, where a metal oxide semiconductor sensor-based electronic olfactory system (MOS array), employed for pathogen bacteria identification based on their volatile organic compound (VOC) characterisation, is assisted by a preliminary separative technique based on gravitational field-flow fractionation (GrFFF). In the integrated system, a preliminary step using GrFFF fractionation of a complex sample provided bacteria-enriched fractions readily available for subsequent MOS array analysis. The MOS array signals were then analysed employing a chemometric approach using principal components analysis (PCA) for a first-data exploration, followed by linear discriminant analysis (LDA) as a classification tool, using the PCA scores as input variables. The ability of the GrFFF-MOS system to distinguish between viable and non-viable cells of the same strain was demonstrated for the first time, yielding 100 % ability of correct prediction. The integrated system was also applied as a proof of concept for multianalyte purposes, for the detection of two bacterial strains (Escherichia coli O157:H7 and Yersinia enterocolitica) simultaneously present in artificially contaminated milk samples, obtaining a 100 % ability of correct prediction. Acquired results show that GrFFF band slicing before MOS array analysis can significantly increase reliability and reproducibility of pathogen bacteria identification based on their VOC production, simplifying the analytical procedure and largely eliminating sample matrix effects. The developed GrFFF-MOS integrated system can be considered a simple straightforward approach for pathogen bacteria identification directly from their food matrix. Graphical abstract An integrated sensing system is presented for pathogen bacteria identification in food, in which field-flow fractionation is exploited to prepare enriched cell fractions prior to their analysis by electronic olfactory system analysis.

Cloning and sequencing of the histidine decarboxylase genes of gram-negative, histamine-producing bacteria and their application in detection and identification of these organisms in fish.

PubMed

Takahashi, Hajime; Kimura, Bon; Yoshikawa, Miwako; Fujii, Tateo

2003-05-01

The use of molecular tools for early and rapid detection of gram-negative histamine-producing bacteria is important for preventing the accumulation of histamine in fish products. To date, no molecular detection or identification system for gram-negative histamine-producing bacteria has been developed. A molecular method that allows the rapid detection of gram-negative histamine producers by PCR and simultaneous differentiation by single-strand conformation polymorphism (SSCP) analysis using the amplification product of the histidine decarboxylase genes (hdc) was developed. A collection of 37 strains of histamine-producing bacteria (8 reference strains from culture collections and 29 isolates from fish) and 470 strains of non-histamine-producing bacteria isolated from fish were tested. Histamine production of bacteria was determined by paper chromatography and confirmed by high-performance liquid chromatography. Among 37 strains of histamine-producing bacteria, all histidine-decarboxylating gram-negative bacteria produced a PCR product, except for a strain of Citrobacter braakii. In contrast, none of the non-histamine-producing strains (470 strains) produced an amplification product. Specificity of the amplification was further confirmed by sequencing the 0.7-kbp amplification product. A phylogenetic tree of the isolates constructed using newly determined sequences of partial hdc was similar to the phylogenetic tree generated from 16S ribosomal DNA sequences. Histamine accumulation occurred when PCR amplification of hdc was positive in all of fish samples tested and the presence of powerful histamine producers was confirmed by subsequent SSCP identification. The potential application of the PCR-SSCP method as a rapid monitoring tool is discussed.

Gold Nanorod-based Photo-PCR System for One-Step, Rapid Detection of Bacteria

PubMed Central

Kim, Jinjoo; Kim, Hansol; Park, Ji Ho; Jon, Sangyong

2017-01-01

The polymerase chain reaction (PCR) has been an essential tool for diagnosis of infectious diseases, but conventional PCR still has some limitations with respect to applications to point-of-care (POC) diagnostic systems that require rapid detection and miniaturization. Here we report a light-based PCR method, termed as photo-PCR, which enables rapid detection of bacteria in a single step. In the photo-PCR system, poly(enthylene glycol)-modified gold nanorods (PEG-GNRs), used as a heat generator, are added into the PCR mixture, which is subsequently periodically irradiated with a 808-nm laser to create thermal cycling. Photo-PCR was able to significantly reduce overall thermal cycling time by integrating bacterial cell lysis and DNA amplification into a single step. Furthermore, when combined with KAPA2G fast polymerase and cooling system, the entire process of bacterial genomic DNA extraction and amplification was further shortened, highlighting the potential of photo-PCR for use in a portable, POC diagnostic system. PMID:29071186

Use of Drosophila S2 Cells as a Model for Studying Ehrlichia chaffeensis Infections▿

PubMed Central

Luce-Fedrow, Alison; Von Ohlen, Tonia; Boyle, Daniel; Ganta, Roman R.; Chapes, Stephen K.

2008-01-01

Ehrlichia chaffeensis is an obligate intracellular bacterium and the causative agent of human monocytic ehrlichiosis. Although this pathogen grows in several mammalian cell lines, no general model for eukaryotic cellular requirements for bacteria replication has yet been proposed. We found that Drosophila S2 cells are permissive for the growth of E. chaffeensis. We saw morulae (aggregates of bacteria) by microscopy, detected the E. chaffeensis 16S rRNA gene by reverse transcriptase PCR, and used immunocytochemistry to detect E. chaffeensis in S2 and mammalian cells. Bacteria grown in S2 cells reinfected mammalian macrophages. S2 cells were made nonpermissive for E. chaffeensis through incubation with lipopolysaccharide. Our results demonstrate that S2 cells are an appropriate system for studying the pathogenesis of E. chaffeensis. The use of a Drosophila system has the potential to serve as a model system for studying Ehrlichia due to its completed genome, ease of genetic manipulation, and the availability of mutants. PMID:18245255

A Review of Green Strategies to Prevent or Mitigate Microbiologically Influenced Corrosion

DTIC Science & Technology

2007-01-01

microbiologically influenced corrosion (MIC) has may provide nutrients for regrowth. been to use oxidising (e.g. chlorine, bromine, ozone ) Videla et al...MPN method: 6cells/cm2 Total bacteria Figure 6. Number of bacteria in biofilm using two detection techniques : fluorescent antibody (FA) and most...nitrite, lation of nitrate- utilising bacteria in the system. nitrous oxide or nitrogen and oxidising sulfide to Hubert et al. (2006) suggested that

Development of a chemiluminescent and bioluminescent system for the detection of bacteria in wastewater effluent

NASA Technical Reports Server (NTRS)

Thomas, R. R.

1975-01-01

Automated chemiluminescent and bioluminescent sensors were developed for continuous monitoring of microbial levels in wastewater effluent. Development of the chemiluminescent system included optimization of reagent concentrations as well as two new techniques which will allow for increased sensitivity and specificity. The optimal reagent concentrations are 0.0025 M luminol and 0.0125 M sodium perborate in 0.75N sodium hydroxide before addition of sample. The methods developed to increase specificity include (1) extraction of porphyrins from bacteria collected in a filter using 0.1N NaOH - 50 percent Ethanol, and (2) use of the specific reaction rate characteristics for the different luminol catalysts. Since reaction times are different for each catalyst, the reaction can be made specific for bacteria by measuring only the light emission from the particular reaction time zone specific for bacteria. Developments of the bioluminescent firefly luciferase system were in the area of flow system design.

Bacteria in atmospheric waters: Detection, characteristics and implications

NASA Astrophysics Data System (ADS)

Hu, Wei; Niu, Hongya; Murata, Kotaro; Wu, Zhijun; Hu, Min; Kojima, Tomoko; Zhang, Daizhou

2018-04-01

In this review paper, we synthesize the current knowledges about bacteria in atmospheric waters, e.g., cloud, fog, rain, and snow, most of which were obtained very recently. First, we briefly describe the importance of bacteria in atmospheric waters, i.e., the essentiality of studying bacteria in atmospheric waters in understanding aerosol-cloud-precipitation-climate interactions in the Earth system. Next, approaches to collect atmospheric water samples for the detection of bacteria and methods to identify the bacteria are summarized and compared. Then the available data on the abundance, viability and community composition of bacteria in atmospheric waters are summarized. The average bacterial concentration in cloud water was usually on the order 104-105 cells mL-1, while that in precipitation on the order 103-104 cells mL-1. Most of the bacteria were viable or metabolically active. Their community composition was highly diverse and differed at various sites. Factors potentially influencing the bacteria, e.g., air pollution levels and sources, meteorological conditions, seasonal effect, and physicochemical properties of atmospheric waters, are described. After that, the implications of bacteria present in atmospheric waters, including their effect on nucleation in clouds, atmospheric chemistry, ecosystems and public health, are briefly discussed. Finally, based on the current knowledges on bacteria in atmospheric waters, which in fact remains largely unknown, we give perspectives that should be paid attention to in future studies.

Asellus aquaticus as a Potential Carrier of Escherichia coli and Other Coliform Bacteria into Drinking Water Distribution Systems

PubMed Central

Christensen, Sarah C. B.; Arvin, Erik; Nissen, Erling; Albrechtsen, Hans-Jørgen

2013-01-01

Individuals of the water louse, Asellus aquaticus, enter drinking water distribution systems in temperate parts of the world, where they establish breeding populations. We analysed populations of surface water A. aquaticus from two ponds for associated faecal indicator bacteria and assessed the risk of A. aquaticus transporting bacteria into distribution systems. Concentrations of up to two E. coli and five total coliforms·mL−1 were measured in the water and 200 E. coli and >240 total coliforms·mL−1 in the sediments of the investigated ponds. Concentrations of A. aquaticus associated bacteria never exceeded three E. coli and six total coliforms·A. aquaticus−1. During exposure to high concentrations of coliforms, concentrations reached 350 coliforms·A. aquaticus−1. A. aquaticus associated E. coli were only detected as long as E. coli were present in the water and sediment. The calculated probability of exceeding drinking water guideline values in non-disinfected systems by intrusion of A. aquaticus was low. Only in scenarios with narrow pipes and low flows, did total coliforms exceed guideline values, implying that the probability of detection by routine monitoring is also low. The study expands the knowledge base for evaluating incidents with presence of coliform indicators in drinking water by showing that intruding A. aquaticus were not important carriers of E. coli or other coliform bacteria even when emerging from faecally contaminated waters. PMID:23455399

Asellus aquaticus as a potential carrier of Escherichia coli and other coliform bacteria into drinking water distribution systems.

PubMed

Christensen, Sarah C B; Arvin, Erik; Nissen, Erling; Albrechtsen, Hans-Jørgen

2013-03-01

Individuals of the water louse, Asellus aquaticus, enter drinking water distribution systems in temperate parts of the world, where they establish breeding populations. We analysed populations of surface water A. aquaticus from two ponds for associated faecal indicator bacteria and assessed the risk of A. aquaticus transporting bacteria into distribution systems. Concentrations of up to two E. coli and five total coliforms·mL-1 were measured in the water and 200 E. coli and >240 total coliforms·mL-1 in the sediments of the investigated ponds. Concentrations of A. aquaticus associated bacteria never exceeded three E. coli and six total coliforms·A. aquaticus-1. During exposure to high concentrations of coliforms, concentrations reached 350 coliforms·A. aquaticus-1. A. aquaticus associated E. coli were only detected as long as E. coli were present in the water and sediment. The calculated probability of exceeding drinking water guideline values in non-disinfected systems by intrusion of A. aquaticus was low. Only in scenarios with narrow pipes and low flows, did total coliforms exceed guideline values, implying that the probability of detection by routine monitoring is also low. The study expands the knowledge base for evaluating incidents with presence of coliform indicators in drinking water by showing that intruding A. aquaticus were not important carriers of E. coli or other coliform bacteria even when emerging from faecally contaminated waters.

Lab-on-a-chip modules for detection of highly pathogenic bacteria: from sample preparation to detection

NASA Astrophysics Data System (ADS)

Julich, S.; Kopinč, R.; Hlawatsch, N.; Moche, C.; Lapanje, A.; Gärtner, C.; Tomaso, H.

2014-05-01

Lab-on-a-chip systems are innovative tools for the detection and identification of microbial pathogens in human and veterinary medicine. The major advantages are small sample volume and a compact design. Several fluidic modules have been developed to transform analytical procedures into miniaturized scale including sampling, sample preparation, target enrichment, and detection procedures. We present evaluation data for single modules that will be integrated in a chip system for the detection of pathogens. A microfluidic chip for purification of nucleic acids was established for cell lysis using magnetic beads. This assay was evaluated with spiked environmental aerosol and swab samples. Bacillus thuringiensis was used as simulant for Bacillus anthracis, which is closely related but non-pathogenic for humans. Stationary PCR and a flow-through PCR chip module were investigated for specific detection of six highly pathogenic bacteria. The conventional PCR assays could be transferred into miniaturized scale using the same temperature/time profile. We could demonstrate that the microfluidic chip modules are suitable for the respective purposes and are promising tools for the detection of bacterial pathogens. Future developments will focus on the integration of these separate modules to an entire lab-on-a-chip system.

Microfluidic Capillaric Circuit for Rapid and Facile Bacteria Detection.

PubMed

Olanrewaju, Ayokunle Oluwafemi; Ng, Andy; DeCorwin-Martin, Philippe; Robillard, Alessandra; Juncker, David

2017-06-20

Urinary tract infections (UTI) are one of the most common bacterial infections and would greatly benefit from a rapid point-of-care diagnostic test. Although significant progress has been made in developing microfluidic systems for nucleic acid and whole bacteria immunoassay tests, their practical application is limited by complex protocols, bulky peripherals, and slow operation. Here we present a microfluidic capillaric circuit (CC) optimized for rapid and automated detection of bacteria in urine. Molds for CCs were constructed using previously established design rules, then 3D-printed and replicated into poly(dimethylsiloxane). CCs autonomously and sequentially performed all liquid delivery steps required for the assay. For efficient bacteria capture, on-the-spot packing of antibody-functionalized microbeads was completed in <20 s followed by autonomous sequential delivery of 100 μL of bacteria sample, biotinylated detection antibodies, fluorescent streptavidin conjugate, and wash buffer for a total volume ≈115 μL. The assay was completed in <7 min. Fluorescence images of the microbead column revealed captured bacteria as bright spots that were easily counted manually or using an automated script for user-independent assay readout. The limit of detection of E. coli in synthetic urine was 1.2 × 10 2 colony-forming-units per mL (CFU/mL), which is well below the clinical diagnostic criterion (>10 5 CFU/mL) for UTI. The self-powered, peripheral-free CC presented here has potential for use in rapid point-of-care UTI screening.

Plant Growth, Antibiotic Uptake, and Prevalence of Antibiotic Resistance in an Endophytic System of Pakchoi under Antibiotic Exposure

PubMed Central

Zhang, Hao; Li, Xunan; Yang, Qingxiang; Sun, Linlin; Yang, Xinxin; Zhou, Mingming; Deng, Rongzhen; Bi, Linqian

2017-01-01

Antibiotic contamination in agroecosystems may cause serious problems, such as the proliferation of various antibiotic resistant bacteria and the spreading of antibiotic resistance genes (ARGs) in the environment or even to human beings. However, it is unclear whether environmental antibiotics, antibiotic resistant bacteria, and ARGs can directly enter into, or occur in, the endophytic systems of plants exposed to pollutants. In this study, a hydroponic experiment exposing pakchoi (Brassica chinensis L.) to tetracycline, cephalexin, and sulfamethoxazole at 50% minimum inhibitory concentration (MIC) levels and MIC levels, respectively, was conducted to explore plant growth, antibiotic uptake, and the development of antibiotic resistance in endophytic systems. The three antibiotics promoted pakchoi growth at 50% MIC values. Target antibiotics at concentrations ranging from 6.9 to 48.1 µg·kg−1 were detected in the treated vegetables. Additionally, the rates of antibiotic-resistant endophytic bacteria to total cultivable endophytic bacteria significantly increased as the antibiotics accumulated in the plants. The detection and quantification of ARGs indicated that four types, tetX, blaCTX-M, and sul1 and sul2, which correspond to tetracycline, cephalexin, and sulfamethoxazole resistance, respectively, were present in the pakchoi endophytic system and increased with the antibiotic concentrations. The results highlight a potential risk of the development and spread of antibiotic resistance in vegetable endophytic systems. PMID:29099753

Plant Growth, Antibiotic Uptake, and Prevalence of Antibiotic Resistance in an Endophytic System of Pakchoi under Antibiotic Exposure.

PubMed

Zhang, Hao; Li, Xunan; Yang, Qingxiang; Sun, Linlin; Yang, Xinxin; Zhou, Mingming; Deng, Rongzhen; Bi, Linqian

2017-11-03

Antibiotic contamination in agroecosystems may cause serious problems, such as the proliferation of various antibiotic resistant bacteria and the spreading of antibiotic resistance genes (ARGs) in the environment or even to human beings. However, it is unclear whether environmental antibiotics, antibiotic resistant bacteria, and ARGs can directly enter into, or occur in, the endophytic systems of plants exposed to pollutants. In this study, a hydroponic experiment exposing pakchoi ( Brassica chinensis L.) to tetracycline, cephalexin, and sulfamethoxazole at 50% minimum inhibitory concentration (MIC) levels and MIC levels, respectively, was conducted to explore plant growth, antibiotic uptake, and the development of antibiotic resistance in endophytic systems. The three antibiotics promoted pakchoi growth at 50% MIC values. Target antibiotics at concentrations ranging from 6.9 to 48.1 µg·kg -1 were detected in the treated vegetables. Additionally, the rates of antibiotic-resistant endophytic bacteria to total cultivable endophytic bacteria significantly increased as the antibiotics accumulated in the plants. The detection and quantification of ARGs indicated that four types, tet X, bla CTX-M , and sul 1 and sul 2, which correspond to tetracycline, cephalexin, and sulfamethoxazole resistance, respectively, were present in the pakchoi endophytic system and increased with the antibiotic concentrations. The results highlight a potential risk of the development and spread of antibiotic resistance in vegetable endophytic systems.

COMPARISON OF MEMBRANE FILTER, MULTIPLE-FERMENTATION-TUBE, AND PRESENCE-ABSENCE TECHNIQUES FOR DETECTING TOTAL COLIFORMS IN SMALL COMMUNITY WATER SYSTEMS

EPA Science Inventory

Methods for detecting total coliform bacteria in drinking water were compared using 1483 different drinking water samples from 15 small community water systems in Vermont and New Hampshire. The methods included the membrane filter (MF) technique, a ten tube fermentation tube tech...

Detection of Biomass in New York City Aerosols: Light Scattering and Optical Fluorescence Techniques

NASA Astrophysics Data System (ADS)

Niebauer, M.; Alimova, A.; Katz, A.; Xu, M.; Rudolph, E.; Steiner, J.; Alfano, R. R.

2005-12-01

Optical spectroscopy is an ideal method for detecting bacteria and spores in real time. Optical fluorescence spectroscopy examination of New York City aerosols is used to quantify the mass of bacteria spores present in air masses collected at 14 liters/minute onto silica fiber filters, and on silica fiber ribbons using an Environmental Beta Attenuation Monitor manufactured by MetOne Instruments configured for the PM2.5 fraction. Dipicolinic acid (DPA), a molecule found primarily in bacterial spores, is the most characteristic component of spores in trial experiments on over 200 collected aerosol samples. DPA is extracted from the spores using a heat bath and chelated with Terbium. The DPA:Tb is detected by measuring its characteristic fluorescence with emission bands at 490, 545 and 585 nm for 270 nm excitation. Light scattering also measures the size distribution for a number of a variety of bacteria - Bacillus subtilis (rod shaped), Staphylococcus aureus (spherical) and Pseudomonas aeruginosa (short rods) establishing that optical techniques satisfactorily distinguish populations based on their variable morphology. Size and morphology are obtained by applying a variation of the Gaussian Ray Approximation theory of anomalous diffraction theory to an analysis of the transmission spectra in the range of 0.4 to 1.0 microns. In test experiments, the refractive index of the inner spore core of Bacillus subtilis decreases from 1.51 to 1.39 while the spore radius enlarges from 0.38 to 0.6 micrometers. Optical determinations are verified by oil-immersion techniques and by scanning electron microscope measurements. Characterization of spores, germinating spore materials, and bacteria is considered vital to tracing bacteria in the environment, for the development of life-detection systems for planetary exploration, monitoring pathogens in environmental systems, and for the preparation of anti-terrorism strategies.

Affinity sensor using 3-aminophenylboronic acid for bacteria detection.

PubMed

Wannapob, Rodtichoti; Kanatharana, Proespichaya; Limbut, Warakorn; Numnuam, Apon; Asawatreratanakul, Punnee; Thammakhet, Chongdee; Thavarungkul, Panote

2010-10-15

Boronic acid that can reversibly bind to diols was used to detect bacteria through its affinity binding reaction with diol-groups on bacterial cell walls. 3-aminophenylboronic acid (3-APBA) was immobilized on a gold electrode via a self-assembled monolayer. The change in capacitance of the sensing surface caused by the binding between 3-APBA and bacteria in a flow system was detected by a potentiostatic step method. Under optimal conditions the linear range of 1.5×10(2)-1.5×10(6) CFU ml(-1) and the detection limit of 1.0×10(2) CFU ml(-1) was obtained. The sensing surface can be regenerated and reused up to 58 times. The method was used for the analysis of bacteria in several types of water, i.e., bottled, well, tap, reservoir and wastewater. Compared with the standard plate count method, the results were within one standard deviation of each other. The proposed method can save both time and cost of analysis. The electrode modified with 3-APBA would also be applicable to the detection of other cis-diol-containing analytes. The concept could be extended to other chemoselective ligands, offering less expensive and more robust affinity sensors for a wide range of compounds. Copyright © 2010 Elsevier B.V. All rights reserved.

Study of Antibacterial Activity and Bacteriology of Latex from Asclepias syriaca L

PubMed Central

McCay, Steven; Mahlberg, Paul

1973-01-01

Whole and fractionated latex of Asclepias syriaca was tested for antimicrobial or growth-promoting activity with 16 genera and species of bacteria. Latex and extracted fractions (distilled water, acetic acid, sodium bicarbonate, sulfuric acid, and ethyl ether) possessed no detectable antimicrobial activity. Comparison of growth curves of selected bacteria incubated with serum and rubber fractions, as well as controls, revealed no detectable inhibition of growth, except for a slight inhibitory response to autoclaved serum. Most bacteria were capable of utilizing latex for a substrate as indicated by the increased growth rate in the exponential phase. The stationary phase was entered simultaneously by both the treated cultures and the controls. Various bacteria cultured in a litmus latex mixture yielded populations which ranged from <104 organisms/ml for Lactobacillus casei, Staphylococcus aureus and Micrococcus lysodeikticus to 1.1 × 1010 organisms/ml for Clostridium acetobutylicum. Whole latex, as well as the serum and rubber fractions, support the growth of various bacteria, but under field conditions there is no evidence for systemic infection of this type of cell by bacteria. PMID:4790590

Tape Cassette Bacteria Detection System

NASA Technical Reports Server (NTRS)

1973-01-01

The design, fabrication, and testing of an automatic bacteria detection system with a zero-g capability and based on the filter-capsule approach is described. This system is intended for monitoring the sterility of regenerated water in a spacecraft. The principle of detection is based on measuring the increase in chemiluminescence produced by the action of bacterial porphyrins (i.e., catalase, cytochromes, etc.) on a luminol-hydrogen peroxide mixture. Since viable as well as nonviable organisms initiate this luminescence, viable organisms are detected by comparing the signal of an incubated water sample with an unincubated control. Higher signals for the former indicate the presence of viable organisms. System features include disposable sealed sterile capsules, each containing a filter membrane, for processing discrete water samples and a tape transport for moving these capsules through a processing sequence which involves sample concentration, nutrient addition, incubation, a 4 Molar Urea wash and reaction with luminol-hydrogen peroxide in front of a photomultiplier tube. Liquids are introduced by means of a syringe needle which pierces a rubber septum contained in the wall of the capsule. Detection thresholds obtained with this unit towards E. coli and S. marcescens assuming a 400 ml water sample are indicated.

Specific Recognition and Detection of MRSA Based on Molecular Probes Comprised of Lytic Phage and Antibody

DTIC Science & Technology

2011-03-29

QCM system was employed to study bacteria-phage interactions. 15. SUBJECT TERMS 16. SECURITY CLASSIFICATION OF: 17. LIMITATION OF ABSTRACT 1 18... QCM -D system (Sweden) was employed to study bacteria-phage interactions. Lytic phages were constructed into hollow spherical particles upon exposure to...a chloroform-water interface. These particles were converted into monolayers and deposited onto QCM -D crystals using Langmuir-Blodgett technique [I

Heterotrophic bacteria in drinking water distribution system: a review.

PubMed

Chowdhury, Shakhawat

2012-10-01

The microbiological quality of drinking water in municipal water distribution systems (WDS) depends on several factors. Free residual chlorine and/or chloramines are typically used to minimize bacterial recontamination and/or regrowth in WDS. Despite such preventive measures, regrowth of heterotrophic (HPC) and opportunistic bacteria in bulk water and biofilms has yet to be controlled completely. No approach has shown complete success in eliminating biofilms or HPC bacteria from bulk water and pipe surfaces. Biofilms can provide shelter for pathogenic bacteria and protect these bacteria from disinfectants. Some HPC bacteria may be associated with aesthetic and non-life threatening diseases. Research to date has achieved important success in understanding occurrence and regrowth of bacteria in bulk water and biofilms in WDS. To achieve comprehensive understanding and to provide efficient control against bacteria regrowth, future research on bacteria regrowth dynamics and their implications is warranted. In this study, a review was performed on the literature published in this area. The findings and limitations of these papers are summarized. Occurrences of bacteria in WDS, factors affecting bacteria regrowth in bulk water and biofilms, bacteria control strategies, sources of nutrients, human health risks from bacterial exposure, modelling of bacteria regrowth and methods of bacteria sampling and detection and quantification are investigated. Advances to date are noted, and future research needs are identified. Finally, research directions are proposed to effectively control HPC and opportunistic bacteria in bulk water and biofilms in WDS.

Combined use of vancomycin-modified Ag-coated magnetic nanoparticles and secondary enhanced nanoparticles for rapid surface-enhanced Raman scattering detection of bacteria.

PubMed

Wang, Chongwen; Gu, Bing; Liu, Qiqi; Pang, Yuanfeng; Xiao, Rui; Wang, Shengqi

2018-01-01

Pathogenic bacteria have always been a significant threat to human health. The detection of pathogens needs to be rapid, accurate, and convenient. We present a sensitive surface-enhanced Raman scattering (SERS) biosensor based on the combination of vancomycin-modified Ag-coated magnetic nanoparticles (Fe 3 O 4 @Ag-Van MNPs) and Au@Ag nanoparticles (NPs) that can effectively capture and discriminate bacterial pathogens from solution. The high-performance Fe 3 O 4 @Ag MNPs were modified with vancomycin and used as bacteria capturer for magnetic separation and enrichment. The modified MNPS were found to exhibit strong affinity with a broad range of Gram-positive and Gram-negative bacteria. After separating and rinsing bacteria, Fe 3 O 4 @Ag-Van MNPs and Au@Ag NPs were synergistically used to construct a very large number of hot spots on bacteria cells, leading to ultrasensitive SERS detection. The dominant merits of our dual enhanced strategy included high bacterial-capture efficiency (>65%) within a wide pH range (pH 3.0-11.0), a short assay time (<30 min), and a low detection limit (5×10 2 cells/mL). Moreover, the spiked tests show that this method is still valid in milk and blood samples. Owing to these capabilities, the combined system enabled the sensitive and specific discrimination of different pathogens in complex solution, as verified by its detection of Gram-positive bacterium Escherichia coli , Gram-positive bacterium Staphylococcus aureus , and methicillin-resistant S. aureus . This method has great potential for field applications in food safety, environmental monitoring, and infectious disease diagnosis.

Combined use of vancomycin-modified Ag-coated magnetic nanoparticles and secondary enhanced nanoparticles for rapid surface-enhanced Raman scattering detection of bacteria

PubMed Central

Pang, Yuanfeng; Xiao, Rui; Wang, Shengqi

2018-01-01

Background Pathogenic bacteria have always been a significant threat to human health. The detection of pathogens needs to be rapid, accurate, and convenient. Methods We present a sensitive surface-enhanced Raman scattering (SERS) biosensor based on the combination of vancomycin-modified Ag-coated magnetic nanoparticles (Fe3O4@Ag-Van MNPs) and Au@Ag nanoparticles (NPs) that can effectively capture and discriminate bacterial pathogens from solution. The high-performance Fe3O4@Ag MNPs were modified with vancomycin and used as bacteria capturer for magnetic separation and enrichment. The modified MNPS were found to exhibit strong affinity with a broad range of Gram-positive and Gram-negative bacteria. After separating and rinsing bacteria, Fe3O4@Ag-Van MNPs and Au@Ag NPs were synergistically used to construct a very large number of hot spots on bacteria cells, leading to ultrasensitive SERS detection. Results The dominant merits of our dual enhanced strategy included high bacterial-capture efficiency (>65%) within a wide pH range (pH 3.0–11.0), a short assay time (<30 min), and a low detection limit (5×102 cells/mL). Moreover, the spiked tests show that this method is still valid in milk and blood samples. Owing to these capabilities, the combined system enabled the sensitive and specific discrimination of different pathogens in complex solution, as verified by its detection of Gram-positive bacterium Escherichia coli, Gram-positive bacterium Staphylococcus aureus, and methicillin-resistant S. aureus. Conclusion This method has great potential for field applications in food safety, environmental monitoring, and infectious disease diagnosis. PMID:29520142

Using Fluorescent Viruses for Detecting Bacteria in Water

NASA Technical Reports Server (NTRS)

Tabacco, Mary Beth; Qian, Xiaohua; Russo, Jaimie A.

2009-01-01

A method of detecting water-borne pathogenic bacteria is based partly on established molecular-recognition and fluorescent-labeling concepts, according to which bacteria of a species of interest are labeled with fluorescent reporter molecules and the bacteria can then be detected by fluorescence spectroscopy. The novelty of the present method lies in the use of bacteriophages (viruses that infect bacteria) to deliver the fluorescent reporter molecules to the bacteria of the species of interest.

Ultrasensitive multiplex optical quantification of bacteria in large samples of biofluids

PubMed Central

Pazos-Perez, Nicolas; Pazos, Elena; Catala, Carme; Mir-Simon, Bernat; Gómez-de Pedro, Sara; Sagales, Juan; Villanueva, Carlos; Vila, Jordi; Soriano, Alex; García de Abajo, F. Javier; Alvarez-Puebla, Ramon A.

2016-01-01

Efficient treatments in bacterial infections require the fast and accurate recognition of pathogens, with concentrations as low as one per milliliter in the case of septicemia. Detecting and quantifying bacteria in such low concentrations is challenging and typically demands cultures of large samples of blood (~1 milliliter) extending over 24–72 hours. This delay seriously compromises the health of patients. Here we demonstrate a fast microorganism optical detection system for the exhaustive identification and quantification of pathogens in volumes of biofluids with clinical relevance (~1 milliliter) in minutes. We drive each type of bacteria to accumulate antibody functionalized SERS-labelled silver nanoparticles. Particle aggregation on the bacteria membranes renders dense arrays of inter-particle gaps in which the Raman signal is exponentially amplified by several orders of magnitude relative to the dispersed particles. This enables a multiplex identification of the microorganisms through the molecule-specific spectral fingerprints. PMID:27364357

A Protein Nanopore-Based Approach for Bacteria Sensing

NASA Astrophysics Data System (ADS)

Apetrei, Aurelia; Ciuca, Andrei; Lee, Jong-kook; Seo, Chang Ho; Park, Yoonkyung; Luchian, Tudor

2016-11-01

We present herein a first proof of concept demonstrating the potential of a protein nanopore-based technique for real-time detection of selected Gram-negative bacteria ( Pseudomonas aeruginosa or Escherichia coli) at a concentration of 1.2 × 108 cfu/mL. The anionic charge on the bacterial outer membrane promotes the electrophoretically driven migration of bacteria towards a single α-hemolysin nanopore isolated in a lipid bilayer, clamped at a negative electric potential, and followed by capture at the nanopore's mouth, which we found to be described according to the classical Kramers' theory. By using a specific antimicrobial peptide as a putative molecular biorecognition element for the bacteria used herein, we suggest that the detection system can combine the natural sensitivity of the nanopore-based sensing techniques with selective biological recognition, in aqueous samples, and highlight the feasibility of the nanopore-based platform to provide portable, sensitive analysis and monitoring of bacterial pathogens.

Probing minority population of antibiotic-resistant bacteria.

PubMed

Huang, Tianxun; Zheng, Yan; Yan, Ya; Yang, Lingling; Yao, Yihui; Zheng, Jiaxin; Wu, Lina; Wang, Xu; Chen, Yuqing; Xing, Jinchun; Yan, Xiaomei

2016-06-15

The evolution and spread of antibiotic-resistant pathogens has become a major threat to public health. Advanced tools are urgently needed to quickly diagnose antibiotic-resistant infections to initiate appropriate treatment. Here we report the development of a highly sensitive flow cytometric method to probe minority population of antibiotic-resistant bacteria via single cell detection. Monoclonal antibody against TEM-1 β-lactamase and Alexa Fluor 488-conjugated secondary antibody were used to selectively label resistant bacteria green, and nucleic acid dye SYTO 62 was used to stain all the bacteria red. A laboratory-built high sensitivity flow cytometer (HSFCM) was applied to simultaneously detect the side scatter and dual-color fluorescence signals of single bacteria. By using E. coli JM109/pUC19 and E. coli JM109 as the model systems for antibiotic-resistant and antibiotic-susceptible bacteria, respectively, as low as 0.1% of antibiotic-resistant bacteria were accurately quantified. By monitoring the dynamic population change of a bacterial culture with the administration of antibiotics, we confirmed that under the antimicrobial pressure, the original low population of antibiotic-resistant bacteria outcompeted susceptible strains and became the dominant population after 5hours of growth. Detection of antibiotic-resistant infection in clinical urine samples was achieved without cultivation, and the bacterial load of susceptible and resistant strains can be faithfully quantified. Overall, the HSFCM-based quantitative method provides a powerful tool for the fundamental studies of antibiotic resistance and holds the potential to provide rapid and precise guidance in clinical therapies. Copyright © 2016 Elsevier B.V. All rights reserved.

Development of a microplate-based fluorescence immunoassay using quantum dot streptavidin conjugates for enumeration of putative marine bacteria, Alteromonas sp., associated with a benthic harpacticoid copepod.

PubMed

Beckman, Erin M; Kawaguchi, Tomohiro; Chandler, G Thomas; Decho, Alan W

2008-12-01

Attached bacteria inhabit the surfaces of many marine animals--a process that may play important roles in the survival and transport through aquatic systems. However, efficient detection of these bacteria has been problematic, especially small aquatic animals such as benthic harpacticoid copepod. Quantum dots (QD) have recently emerged as a significant tool in immunofluorescence detection because of their unique properties compared to other fluorescent probes. In the present study, a polyclonal antibody was raised against the Gram-negative marine bacterium, Alteromonas sp. A microplate-based immunofluorescence bioassay using QD strepavidin conjugates was developed for quantifying putative Alteromonas sp. cells located on the surfaces of a marine harpacticoid copepod, Microarthridion littorale. The number of attached Alteromonas sp. was estimated to be 10(2)+/-8 CFU using this method. The QD approach, coupled to a microplate assay can potentially provide an efficient and accurate method for rapidly detecting multiple bacteria species attached to small invertebrate animals because of their unique excitation and emission characteristics.

Viable bacteria associated with red blood cells and plasma in freshly drawn blood donations.

PubMed

Damgaard, Christian; Magnussen, Karin; Enevold, Christian; Nilsson, Martin; Tolker-Nielsen, Tim; Holmstrup, Palle; Nielsen, Claus Henrik

2015-01-01

Infection remains a leading cause of post-transfusion mortality and morbidity. Bacterial contamination is, however, detected in less than 0.1% of blood units tested. The aim of the study was to identify viable bacteria in standard blood-pack units, with particular focus on bacteria from the oral cavity, and to determine the distribution of bacteria revealed in plasma and in the red blood cell (RBC)-fraction. Cross-sectional study. Blood were separated into plasma and RBC-suspensions, which were incubated anaerobically or aerobically for 7 days on trypticase soy blood agar (TSA) or blue lactose plates. For identification colony PCR was performed using primers targeting 16S rDNA. Blood donors attending Capital Region Blood Bank, Copenhagen University Hospital, Rigshospitalet, Hvidovre, Denmark, October 29th to December 10th 2013. 60 donors (≥50 years old), self-reported medically healthy. Bacterial growth was observed on plates inoculated with plasma or RBCs from 62% of the blood donations. Growth was evident in 21 (35%) of 60 RBC-fractions and in 32 (53%) of 60 plasma-fractions versus 8 of 60 negative controls (p = 0.005 and p = 2.6x10-6, respectively). Propionibacterium acnes was found in 23% of the donations, and Staphylococcus epidermidis in 38%. The majority of bacteria identified in the present study were either facultative anaerobic (59.5%) or anaerobic (27.8%) species, which are not likely to be detected during current routine screening. Viable bacteria are present in blood from donors self-reported as medically healthy, indicating that conventional test systems employed by blood banks insufficiently detect bacteria in plasma. Further investigation is needed to determine whether routine testing for anaerobic bacteria and testing of RBC-fractions for adherent bacteria should be recommended.

Coliform Bacteria Monitoring in Fish Systems: Current Practices in Public Aquaria.

PubMed

Culpepper, Erin E; Clayton, Leigh A; Hadfield, Catherine A; Arnold, Jill E; Bourbon, Holly M

2016-06-01

Public aquaria evaluate coliform indicator bacteria levels in fish systems, but the purpose of testing, testing methods, and management responses are not standardized, unlike with the coliform bacteria testing for marine mammal enclosures required by the U.S. Department of Agriculture. An online survey was sent to selected aquaria to document current testing and management practices in fish systems without marine mammals. The information collected included indicator bacteria species, the size and type of systems monitored, the primary purpose of testing, sampling frequency, test methods, the criteria for interpreting results, corrective actions, and management changes to limit human exposure. Of the 25 institutions to which surveys were sent, 19 (76%) responded. Fourteen reported testing for fecal indicator bacteria in fish systems. The most commonly tested indicator species were total (86%) and fecal (79%) coliform bacteria, which were detected by means of the membrane filtration method (64%). Multiple types and sizes of systems were tested, and the guidelines for testing and corrective actions were highly variable. Only three institutions performed additional tests to confirm the identification of indicator organisms. The results from this study can be used to compare bacterial monitoring practices and protocols in fish systems, as an aid to discussions relating to the accuracy and reliability of test results, and to help implement appropriate management responses. Received August 23, 2015; accepted December 29, 2015.

Examination and characterization of distribution system biofilms.

PubMed Central

LeChevallier, M W; Babcock, T M; Lee, R G

1987-01-01

Investigations concerning the role of distribution system biofilms on water quality were conducted at a drinking water utility in New Jersey. The utility experienced long-term bacteriological problems in the distribution system, while treatment plant effluents were uniformly negative for coliform bacteria. Results of a monitoring program showed increased coliform levels as the water moved from the treatment plant through the distribution system. Increased coliform densities could not be accounted for by growth of the cells in the water column alone. Identification of coliform bacteria showed that species diversity increased as water flowed through the study area. All materials in the distribution system had high densities of heterotrophic plate count bacteria, while high levels of coliforms were detected only in iron tubercles. Coliform bacteria with the same biochemical profile were found both in distribution system biofilms and in the water column. Assimilable organic carbon determinations showed that carbon levels declined as water flowed through the study area. Maintenance of a 1.0-mg/liter free chlorine residual was insufficient to control coliform occurrences. Flushing and pigging the study area was not an effective control for coliform occurrences in that section. Because coliform bacteria growing in distribution system biofilms may mask the presence of indicator organisms resulting from a true breakdown of treatment barriers, the report recommends that efforts continue to find methods to control growth of coliform bacteria in pipeline biofilms. Images PMID:3435140

Bacterial nitrification in chloraminated water supplies.

PubMed Central

Cunliffe, D A

1991-01-01

Nitrifying bacteria were detected in 64% of samples collected from five chloraminated water supplies in South Australia and in 20.7% of samples that contained more than 5.0 mg of monochloramine per liter. Laboratory experiments confirmed that nitrifying bacteria are relatively resistant to the disinfectant. Increased numbers of the bacteria were associated with accelerated decays of monochloramine within distribution systems. The combination of increased concentrations of oxidized nitrogen with decreased total chlorine residuals can be used as a rapid indicator of bacterial nitrification. PMID:1781698

Method of Detecting Coliform Bacteria and Escherichia Coli Bacteria from Reflected Light

NASA Technical Reports Server (NTRS)

Vincent, Robert (Inventor)

2013-01-01

The present invention relates to a method of detecting coliform bacteria in water from reflected light and a method of detecting Eschericha Coli bacteria in water from reflected light, and also includes devices for the measurement, calculation and transmission of data relating to that method.

Impact of the reusing of food manufacturing wastewater for irrigation in a closed system on the microbiological quality of the food crops.

PubMed

Beneduce, Luciano; Gatta, Giuseppe; Bevilacqua, Antonio; Libutti, Angela; Tarantino, Emanuele; Bellucci, Micol; Troiano, Eleonora; Spano, Giuseppe

2017-11-02

In order to evaluate if the reuse of food industry treated wastewater is compatible for irrigation of food crops, without increased health risk, in the present study a cropping system, in which ground water and treated wastewater were used for irrigation of tomato and broccoli, during consecutive crop seasons was monitored. Water, crop environment and final products were monitored for microbial indicators and pathogenic bacteria, by conventional and molecular methods. The microbial quality of the irrigation waters influenced sporadically the presence of microbial indicators in soil. No water sample was found positive for pathogenic bacteria, independently from the source. Salmonella spp. and Listeria monocytogenes were detected in soil samples, independently from the irrigation water source. No pathogen was found to contaminate tomato plants, while Listeria monocytogenes and E. coli O157:H7 were detected on broccoli plant, but when final produce were harvested, no pathogen was detected on edible part. The level of microbial indicators and detection of pathogenic bacteria in field and plant was not dependent upon wastewater used. Our results, suggest that reuse of food industry wastewater for irrigation of agricultural crop can be applied without significant increase of potential health risk related to microbial quality. Copyright © 2017 Elsevier B.V. All rights reserved.

High-volume extraction of nucleic acids by magnetic bead technology for ultrasensitive detection of bacteria in blood components.

PubMed

Störmer, Melanie; Kleesiek, Knut; Dreier, Jens

2007-01-01

Nucleic acid isolation, the most technically demanding and laborious procedure performed in molecular diagnostics, harbors the potential for improvements in automation. A recent development is the use of magnetic beads covered with nucleic acid-binding matrices. We adapted this technology with a broad-range 23S rRNA real-time reverse transcription (RT)-PCR assay for fast and sensitive detection of bacterial contamination of blood products. We investigated different protocols for an automated high-volume extraction method based on magnetic-separation technology for the extraction of bacterial nucleic acids from platelet concentrates (PCs). We added 2 model bacteria, Staphylococcus epidermidis and Escherichia coli, to a single pool of apheresis-derived, single-donor platelets and assayed the PCs by real-time RT-PCR analysis with an improved primer-probe system and locked nucleic acid technology. Co-amplification of human beta(2)-microglobulin mRNA served as an internal control (IC). We used probit analysis to calculate the minimum concentration of bacteria that would be detected with 95% confidence. For automated magnetic bead-based extraction technology with the real-time RT-PCR, the 95% detection limit was 29 x 10(3) colony-forming units (CFU)/L for S. epidermidis and 22 x 10(3) CFU/L for E. coli. No false-positive results occurred, either due to nucleic acid contamination of reagents or externally during testing of 1030 PCs. High-volume nucleic acid extraction improved the detection limit of the assay. The improvement of the primer-probe system and the integration of an IC make the RT-PCR assay appropriate for bacteria screening of platelets.

Bacterial Signaling to the Nervous System through Toxins and Metabolites.

PubMed

Yang, Nicole J; Chiu, Isaac M

2017-03-10

Mammalian hosts interface intimately with commensal and pathogenic bacteria. It is increasingly clear that molecular interactions between the nervous system and microbes contribute to health and disease. Both commensal and pathogenic bacteria are capable of producing molecules that act on neurons and affect essential aspects of host physiology. Here we highlight several classes of physiologically important molecular interactions that occur between bacteria and the nervous system. First, clostridial neurotoxins block neurotransmission to or from neurons by targeting the SNARE complex, causing the characteristic paralyses of botulism and tetanus during bacterial infection. Second, peripheral sensory neurons-olfactory chemosensory neurons and nociceptor sensory neurons-detect bacterial toxins, formyl peptides, and lipopolysaccharides through distinct molecular mechanisms to elicit smell and pain. Bacteria also damage the central nervous system through toxins that target the brain during infection. Finally, the gut microbiota produces molecules that act on enteric neurons to influence gastrointestinal motility, and metabolites that stimulate the "gut-brain axis" to alter neural circuits, autonomic function, and higher-order brain function and behavior. Furthering the mechanistic and molecular understanding of how bacteria affect the nervous system may uncover potential strategies for modulating neural function and treating neurological diseases. Copyright © 2017 Elsevier Ltd. All rights reserved.

Colonization of plants by human pathogenic bacteria in the course of organic vegetable production.

PubMed

Hofmann, Andreas; Fischer, Doreen; Hartmann, Anton; Schmid, Michael

2014-01-01

In recent years, increasing numbers of outbreaks caused by the consumption of vegetables contaminated with human pathogenic bacteria were reported. The application of organic fertilizers during vegetable production is one of the possible reasons for contamination with those pathogens. In this study laboratory experiments in axenic and soil systems following common practices in organic farming were conducted to identify the minimal dose needed for bacterial colonization of plants and to identify possible factors like bacterial species or serovariation, plant species or organic fertilizer types used, influencing the success of plant colonization by human pathogenic bacteria. Spinach and corn salad were chosen as model plants and were inoculated with different concentrations of Salmonella enterica sv. Weltevreden, Listeria monocytogenes sv. 4b and EGD-E sv. 1/2a either directly (axenic system) or via agricultural soil amended with spiked organic fertilizers (soil system). In addition to PCR- and culture-based detection methods, fluorescence in situ hybridization (FISH) was applied in order to localize bacteria on or in plant tissues. Our results demonstrate that shoots were colonized by the pathogenic bacteria at inoculation doses as low as 4 × 10 CFU/ml in the axenic system or 4 × 10(5) CFU/g in the soil system. In addition, plant species dependent effects were observed. Spinach was colonized more often and at lower inoculation doses compared to corn salad. Differential colonization sites on roots, depending on the plant species could be detected using FISH-CLSM analysis. Furthermore, the transfer of pathogenic bacteria to plants via organic fertilizers was observed more often and at lower initial inoculation doses when fertilization was performed with inoculated slurry compared to inoculated manure. Finally, it could be shown that by introducing a simple washing step, the bacterial contamination was reduced in most cases or even was removed completely in some cases.

Colonization of plants by human pathogenic bacteria in the course of organic vegetable production

PubMed Central

Hofmann, Andreas; Fischer, Doreen; Hartmann, Anton; Schmid, Michael

2014-01-01

In recent years, increasing numbers of outbreaks caused by the consumption of vegetables contaminated with human pathogenic bacteria were reported. The application of organic fertilizers during vegetable production is one of the possible reasons for contamination with those pathogens. In this study laboratory experiments in axenic and soil systems following common practices in organic farming were conducted to identify the minimal dose needed for bacterial colonization of plants and to identify possible factors like bacterial species or serovariation, plant species or organic fertilizer types used, influencing the success of plant colonization by human pathogenic bacteria. Spinach and corn salad were chosen as model plants and were inoculated with different concentrations of Salmonella enterica sv. Weltevreden, Listeria monocytogenes sv. 4b and EGD-E sv. 1/2a either directly (axenic system) or via agricultural soil amended with spiked organic fertilizers (soil system). In addition to PCR- and culture-based detection methods, fluorescence in situ hybridization (FISH) was applied in order to localize bacteria on or in plant tissues. Our results demonstrate that shoots were colonized by the pathogenic bacteria at inoculation doses as low as 4 × 10 CFU/ml in the axenic system or 4 × 105 CFU/g in the soil system. In addition, plant species dependent effects were observed. Spinach was colonized more often and at lower inoculation doses compared to corn salad. Differential colonization sites on roots, depending on the plant species could be detected using FISH-CLSM analysis. Furthermore, the transfer of pathogenic bacteria to plants via organic fertilizers was observed more often and at lower initial inoculation doses when fertilization was performed with inoculated slurry compared to inoculated manure. Finally, it could be shown that by introducing a simple washing step, the bacterial contamination was reduced in most cases or even was removed completely in some cases. PMID:24829562

Label and label-free based surface-enhanced Raman scattering for pathogen bacteria detection: A review.

PubMed

Liu, Yu; Zhou, Haibo; Hu, Ziwei; Yu, Guangxia; Yang, Danting; Zhao, Jinshun

2017-08-15

Rapid, accurate detection of pathogen bacteria is a highly topical research area for the sake of food safety and public health. Surface-enhanced Raman scattering (SERS) is being considered as a powerful and attractive technique for pathogen bacteria detection, due to its sensitivity, high speed, comparatively low cost, multiplexing ability and portability. This contribution aims to give a comprehensive overview of SERS as a technique for rapid detection of pathogen bacteria based on label and label-free strategies. A brief tutorial on SERS is given first of all. Then we summarize the recent trends and developments of label and label-free based SERS applied to detection of pathogen bacteria, including the relatively complete interpretation of SERS spectra. In addition, multifunctional SERS platforms for pathogen bacteria in matrix are discussed as well. Furthermore, an outlook of the work done and a perspective on the future directions of SERS as a reliable tool for real-time pathogen bacteria detection are given. Copyright © 2017 Elsevier B.V. All rights reserved.

Detection of selected periodontal bacteria in preschool children affected by early childhood caries.

PubMed

Pantuckova, Pavla; Bartosova, Michaela; Broukal, Zdenek; Kukletova, Martina; Holla, Lydie Izakovicova

2016-11-01

The aim of this study was to compare the detection frequency of periodontal bacteria in dental plaque in children with early childhood caries (ECC) with and without gingival inflammation. A convenience sample of 25 preschool children (mean age 3.61 years, SD 1.42) was recruited. Dental plaque was taken from periodontal areas with and without visible signs of inflammation and processed using the StomaGene® (Protean s.r.o. Czech Republic) and ParoCheck® 20 (Greiner Bio-one GmbH, Germany) detection kits. The two sample t tests between percents for differences between inflammatory and healthy sites and kappa statistics for the agreement of both systems were used. At the inflammatory sites, Porphyromonas gingivalis and Aggregatibacter actinomycetemcomitans were significantly more frequently detected by StomaGene® while Fusobacterium nucleatum, A. actinomycetemcomitans, Tanarella forsythia and Prevotella intermedia were significantly more frequently identified by ParoCheck® test. The agreement between the two detection systems was substantial for A. actinomycetemcomitans and F. nucleatum in the samples collected from inflamed sites and only for F. nucleatum from clinically healthy sites. Therefore, we recommend that the same system should be used when the same patient is examined repeatedly.

Detection of Biological Pathogens Using Multiple Wireless Magnetoelastic Biosensors

NASA Astrophysics Data System (ADS)

Shen, Wen

A number of recent, high-profile incidences of food-borne illness spreading through the food supply and the use of anthrax by terrorists after the September 11, 2001 attacks have demonstrated the need for new technologies that can rapidly detect the presence of biological pathogens. A bevy of biosensors show excellent detection sensitivity and specificity. However, false positive and false negative signals remain one of the primary reasons that many of these newly developed biosensors have not found application in the marketplace. The research described in this dissertation focuses on developing a free-standing magnetoelastic based bio-sensing system using a pulse method. This method allows fast detection, eliminates the bias magnetic field that is necessary in current methods, makes the system more simply and suitable for in-field detection. This system has two pairs of transformer coils, where a measurement sensor and a control sensor can be put in each pair of coils. The control sensor is used to compensate for environmental variables. The effect of pulse power on the performance of the magnetoelastic sensors in the pulse system is studied. The system is found to have excellent stability, good detection repeatability when used with multiple sensors. This research has investigated and demonstrated a multiple sensors approach. Because it will involve the simultaneous measurement of many sensors, it will significantly reduce problems encountered with false positive indications. The positioning and interference of sensors are investigated. By adding a multi-channel structure to the pulse detection system, the effect of sensor interference is minimized. The result of the repeatability test shows that the standard deviation when measuring three 1 mm magnetoelastic sensors is around 500 Hz, which is smaller than the minimum requirement for actual spores/bacteria detection. Magnetoelastic sensors immobilized with JRB7 phages and E2 phages have been used to specifically detect Bacillus anthracis spores and Salmonella typhimurium bacteria. The real-time monitoring of the detection of B. anthracis spores in a flowing system was performed using 2 mm sensors and 1 mm sensors. The detection of S. typhimurium in air has been performed using the pulse based system with both single and grouped sensors. Because grouped sensor detection involves the simultaneous measurement of many sensors, statistical evaluation shows that it can significantly reduce problems encountered with false positive indications. This method has been implemented in an investigation of a method that allows direct detection of S. typhimurium on cantaloupe surfaces. It has been demonstrated that multiple E2 phage based magnetoelastic sensors are able to detect Salmonella directly on fresh cantaloupe surfaces. Confirmation of the spore or bacteria binding to the sensor surfaces was achieved through SEM study of the sensor surfaces.

Integrated site-specific quantification of faecal bacteria and detection of DNA markers in faecal contamination source tracking as a microbial risk tracking tool in urban Lake ecosystems

NASA Astrophysics Data System (ADS)

Donde, Oscar Omondi; Tian, Cuicui; Xiao, Bangding

2017-11-01

The presence of feacal-derived pathogens in water is responsible for several infectious diseases and deaths worldwide. As a solution, sources of fecal pollution in waters must be accurately assessed, properly determined and strictly controlled. However, the exercise has remained challenging due to the existing overlapping characteristics by different members of faecal coliform bacteria and the inadequacy of information pertaining to the contribution of seasonality and weather condition on tracking the possible sources of pollution. There are continued efforts to improve the Faecal Contamination Source Tracking (FCST) techniques such as Microbial Source Tracking (MST). This study aimed to make contribution to MST by evaluating the efficacy of combining site specific quantification of faecal contamination indicator bacteria and detection of DNA markers while accounting for seasonality and weather conditions' effects in tracking the major sources of faecal contamination in a freshwater system (Donghu Lake, China). The results showed that the use of cyd gene in addition to lacZ and uidA genes differentiates E. coli from other closely related faecal bacteria. The use of selective media increases the pollution source tracking accuracy. BSA addition boosts PCR detection and increases FCST efficiency. Seasonality and weather variability also influence the detection limit for DNA markers.

Performance of two blood culture systems to detect anaerobic bacteria. Is there any difference?

PubMed

Mueller-Premru, Manica; Jeverica, Samo; Papst, Lea; Nagy, Elisabeth

2017-06-01

We studied the performance characteristics of two blood culture (BC) bottles/systems, (i) BacT/ALERT-FN Plus/3D (bioMérieux, Marcy l'Étoile, France) and (ii) BACTEC-Lytic/9000 (Becton Dickinson, Sparks, USA) for detection of growth and time-to-positivity (TTP) against a balanced and diverse collection of anaerobic bacterial strains (n = 48) that included reference strains (n = 19) and clinical isolates (n = 29) of 32 species (15 Gram-negative and 17 Gram-positive). Standard suspension of bacteria was inoculated to each bottle in duplicates and incubated in the corresponding system. Overall, 62.5% (n = 30) of strains were detected by both BC bottle types. Comparing the two, 70.8% (n = 34) and 79.2% (n = 38) of strains were detected by BacT/ALERT-FN Plus and BACTEC-Lytic bottles, respectively (p = 0.38). Among Gram-negative anaerobes (n = 25) the detection rate was 76.0% (n = 19) vs. 92.0% (n = 23) (p = 0.22), respectively. Among Gram-positive anaerobes (n = 23) the detection rate was 65.2% (n = 15) in both bottles (p = 1). The average TTP per bottle was calculated only for the strains detected by both systems (n = 30) and was 40.85 h and 28.08 h for BacT/ALERT-FN Plus and BACTEC-Lytic, respectively (p < 0.001). The mean difference was 12.76 h (95% CI: 6.21-19-31 h). Six anaerobic strains were not detected by any system, including Gram-negative Porphyromonas gingivalis, and five Gram-positive strains: Finegoldia magna, Peptostreptococcus anaerobius, Propionibacterium acnes, Clostridium novyi and Clostridium clostridioforme. Furthermore, Eggerthella lenta and Prevotella bivia were detected only by BacT/ALERT-FN Plus, while Prevotella disiens and Prevotella intermedia were detected only by BACTEC-Lytic bottles. There were no major differences in detection rate among clinical and reference strains. Anaerobic bacteria represent a minority of BC isolates, however, far from ideal detection rate was observed in this study for both tested bottle/system combinations. Nevertheless, in those cases where both gave positive signal, BACTEC-Lytic was superior to BacT/ALERT FN Plus with 12.76 h shorter mean TTP. Improvements of media in blood culture bottles available for detection of anaerobes are warranted. Copyright © 2017 The Authors. Published by Elsevier Ltd.. All rights reserved.

Machine Reading for Extraction of Bacteria and Habitat Taxonomies

PubMed Central

Kordjamshidi, Parisa; Massa, Wouter; Provoost, Thomas; Moens, Marie-Francine

2015-01-01

There is a vast amount of scientific literature available from various resources such as the internet. Automating the extraction of knowledge from these resources is very helpful for biologists to easily access this information. This paper presents a system to extract the bacteria and their habitats, as well as the relations between them. We investigate to what extent current techniques are suited for this task and test a variety of models in this regard. We detect entities in a biological text and map the habitats into a given taxonomy. Our model uses a linear chain Conditional Random Field (CRF). For the prediction of relations between the entities, a model based on logistic regression is built. Designing a system upon these techniques, we explore several improvements for both the generation and selection of good candidates. One contribution to this lies in the extended exibility of our ontology mapper that uses an advanced boundary detection and assigns the taxonomy elements to the detected habitats. Furthermore, we discover value in the combination of several distinct candidate generation rules. Using these techniques, we show results that are significantly improving upon the state of art for the BioNLP Bacteria Biotopes task. PMID:27077141

Rapid detection of microbes in the dialysis solution by the microcolony fluorescence staining method (Millflex quantum).

PubMed

Osono, Eiichi; Kobayashi, Eiko; Inoue, Yuki; Honda, Kazumi; Kumagai, Takuya; Negishi, Hideki; Okamatsu, Kentaro; Ichimura, Kyoko; Kamano, Chisako; Suzuki, Fumi; Norose, Yoshihiko; Takahashi, Megumi; Takaku, Shun; Fujioka, Noriaki; Hayama, Naoaki; Takizawa, Hideaki

2014-01-01

A chemiluminescence system, Milliflex Quantum (MFQ), to detect microcolonies, has been used in the pharmaceutical field. In this study, we investigated aquatic bacteria in hemodialysis solutions sampled from bioburden areas in 4 dialysis faculties. Using MFQ, microcolonies could be detected after a short incubation period. The colony count detected with MFQ after a 48-hour incubation was 92% ± 39%, compared to that after the conventionally used 7-14-day incubation period; in addition, the results also showed a linear correlation. Moreover, MFQ-based analysis allowed the visualization of damaged cells and of the high density due to the excessive amount of bacteria. These results suggested that MFQ had adequate sensitivity to detect microbacteria in dialysis solutions, and it was useful for validating the conditions of conventional culture methods.

Hydrogen sulfide production from cysteine and homocysteine by periodontal and oral bacteria.

PubMed

Yoshida, Akihiro; Yoshimura, Mamiko; Ohara, Naoya; Yoshimura, Shigeru; Nagashima, Shiori; Takehara, Tadamichi; Nakayama, Koji

2009-11-01

Hydrogen sulfide is one of the predominant volatile sulfur compounds (VSCs) produced by oral bacteria. This study developed and evaluated a system for detecting hydrogen sulfide production by oral bacteria. L-methionine-alpha-deamino-gamma-mercaptomethane-lyase (METase) and beta carbon-sulfur (beta C-S) lyase were used to degrade homocysteine and cysteine, respectively, to produce hydrogen sulfide. Enzymatic reactions resulting in hydrogen sulfide production were assayed by reaction with bismuth trichloride, which forms a black precipitate when mixed with hydrogen sulfide. The enzymatic activities of various oral bacteria that result in hydrogen sulfide production and the capacity of bacteria from periodontal sites to form hydrogen sulfide in reaction mixtures containing L-cysteine or DL-homocysteine were assayed. With L-cysteine as the substrate, Streptococcus anginosus FW73 produced the most hydrogen sulfide, whereas Porphyromonas gingivalis American Type Culture Collection (ATCC) 33277 and W83 and Fusobacterium nucleatum ATCC 10953 produced approximately 35% of the amount produced by the P. gingivalis strains. Finally, the hydrogen sulfide found in subgingival plaque was analyzed. Using bismuth trichloride, the hydrogen sulfide produced by oral bacteria was visually detectable as a black precipitate. Hydrogen sulfide production by oral bacteria was easily analyzed using bismuth trichloride. However, further innovation is required for practical use.

9 CFR 113.26 - Detection of viable bacteria and fungi except in live vaccine.

Code of Federal Regulations, 2011 CFR

2011-01-01

... 9 Animals and Animal Products 1 2011-01-01 2011-01-01 false Detection of viable bacteria and fungi... VECTORS STANDARD REQUIREMENTS Standard Procedures § 113.26 Detection of viable bacteria and fungi except... required to be free of viable bacteria and fungi, they shall also be tested as prescribed in this section...

9 CFR 113.26 - Detection of viable bacteria and fungi except in live vaccine.

Code of Federal Regulations, 2010 CFR

2010-01-01

... 9 Animals and Animal Products 1 2010-01-01 2010-01-01 false Detection of viable bacteria and fungi... VECTORS STANDARD REQUIREMENTS Standard Procedures § 113.26 Detection of viable bacteria and fungi except... required to be free of viable bacteria and fungi, they shall also be tested as prescribed in this section...

In vitro model of production of antibodies; a new approach to reveal the presence of key bacteria in polymicrobial environments.

PubMed

Wu, Chongcong; Nakka, Sravya; Mansouri, Sepahdar; Bengtsson, Torbjörn; Nayeri, Tayeb; Nayeri, Fariba

2016-09-09

There is a rapid emergence of multiple resistant gram-negative bacteria due to overuse of antibiotics in the treatment of infections. Biofilms consist of polymicrobial communities that survive the host's defense system. The key bacteria in biofilms are slow growing and support an attachment and rapid growth of other microorganisms. Current antimicrobial strategies often fail due to poor diagnosis of key pathogens in biofilms. The study aims to develop anti-bacterial human antibodies in vitro from patients who had recently undergone a systemic infection by pathogenic bacteria and to use these antibodies as a tool for detecting bacteria in biofilms. Lymphocytes were separated from whole blood of patients (n = 10) and stimulated with heat-killed bacteria to produce antibodies in vitro. The specificity of antibodies in recognizing the bacteria against which they were directed was evaluated by surface plasmon resonance system (SPR) and electron microscopy. The ulcer secretions from patients with chronic and acute leg ulcers and healthy controls were analyzed by the SPR system and the results were compared with culture studies. The produced antibodies recognized bacteria with high sensitivity (SPR). The antibodies against Enterococcus fecalis bound specifically to the microorganism in a bacterial co-culture that was visualized by electron microscopy. In the present work, a method for producing specific antibodies against bacteria is introduced to recognize bacterial components in body fluids of patients suffering from pathogenic biofilms. This diagnostic technique may be most useful in clinical microbiology and in the choice of antibiotics in the treatment of serious infections.

Assessing the origin of bacteria in tap water and distribution system in an unchlorinated drinking water system by SourceTracker using microbial community fingerprints.

PubMed

Liu, Gang; Zhang, Ya; van der Mark, Ed; Magic-Knezev, Aleksandra; Pinto, Ameet; van den Bogert, Bartholomeus; Liu, Wentso; van der Meer, Walter; Medema, Gertjan

2018-07-01

The general consensus is that the abundance of tap water bacteria is greatly influenced by water purification and distribution. Those bacteria that are released from biofilm in the distribution system are especially considered as the major potential risk for drinking water bio-safety. For the first time, this full-scale study has captured and identified the proportional contribution of the source water, treated water, and distribution system in shaping the tap water bacterial community based on their microbial community fingerprints using the Bayesian "SourceTracker" method. The bacterial community profiles and diversity analyses illustrated that the water purification process shaped the community of planktonic and suspended particle-associated bacteria in treated water. The bacterial communities associated with suspended particles, loose deposits, and biofilm were similar to each other, while the community of tap water planktonic bacteria varied across different locations in distribution system. The microbial source tracking results showed that there was not a detectable contribution of source water to bacterial community in the tap water and distribution system. The planktonic bacteria in the treated water was the major contributor to planktonic bacteria in the tap water (17.7-54.1%). The particle-associated bacterial community in the treated water seeded the bacterial community associated with loose deposits (24.9-32.7%) and biofilm (37.8-43.8%) in the distribution system. In return, the loose deposits and biofilm showed a significant influence on tap water planktonic and particle-associated bacteria, which were location dependent and influenced by hydraulic changes. This was revealed by the increased contribution of loose deposits to tap water planktonic bacteria (from 2.5% to 38.0%) and an increased contribution of biofilm to tap water particle-associated bacteria (from 5.9% to 19.7%) caused by possible hydraulic disturbance from proximal to distal regions. Therefore, our findings indicate that the tap water bacteria could possibly be managed by selecting and operating the purification process properly and cleaning the distribution system effectively. Copyright © 2018 Elsevier Ltd. All rights reserved.

Distribution of Escherichia Coli as Soil Pollutant around Antang Landfills

NASA Astrophysics Data System (ADS)

Artiningsih, Andi; Zubair, Hazairin; Imran, A. M.; Widodo, Sri

2018-03-01

Tamangapa Antang Landfill locates around the residential area and faces an air and water pollution due to an open dumping system in its operation. The system arises a potential pollution in air, water and soil. Sampling was done surround the landfill in two parts, parallel and perpendicular to the ground water flow. This study shows the abundance of E. coli bacteria in soil around the Antang Landfills at depth of 10 to 20 cm (93x105 cfu/gr of soil) in the direction of groundwater flow. While in other locations the E. coli bacteria is not detected. The abundance of E. coli bacteria is a conjunction factor from landfill and human activities surround the area. The absence of E. coli bacteria in other location highly interpreted that the landfill is the major contributor of pollutant.

Rapid detection of bacteria in foods and biological fluids

NASA Technical Reports Server (NTRS)

Fealey, R. D.; Renner, W.

1973-01-01

Simple and inexpensive apparatus, called "redox monitoring cell," rapidly detects presence of bacteria. Bacteria is detected by measuring drop in oxygen content in test solution. Apparatus consists of vial with two specially designed electrodes connected to sensitive voltmeter.

Method and automated apparatus for detecting coliform organisms

NASA Technical Reports Server (NTRS)

Dill, W. P.; Taylor, R. E.; Jeffers, E. L. (Inventor)

1980-01-01

Method and automated apparatus are disclosed for determining the time of detection of metabolically produced hydrogen by coliform bacteria cultured in an electroanalytical cell from the time the cell is inoculated with the bacteria. The detection time data provides bacteria concentration values. The apparatus is sequenced and controlled by a digital computer to discharge a spent sample, clean and sterilize the culture cell, provide a bacteria nutrient into the cell, control the temperature of the nutrient, inoculate the nutrient with a bacteria sample, measures the electrical potential difference produced by the cell, and measures the time of detection from inoculation.

Cloud-enabled microscopy and droplet microfluidic platform for specific detection of Escherichia coli in water.

PubMed

Golberg, Alexander; Linshiz, Gregory; Kravets, Ilia; Stawski, Nina; Hillson, Nathan J; Yarmush, Martin L; Marks, Robert S; Konry, Tania

2014-01-01

We report an all-in-one platform - ScanDrop - for the rapid and specific capture, detection, and identification of bacteria in drinking water. The ScanDrop platform integrates droplet microfluidics, a portable imaging system, and cloud-based control software and data storage. The cloud-based control software and data storage enables robotic image acquisition, remote image processing, and rapid data sharing. These features form a "cloud" network for water quality monitoring. We have demonstrated the capability of ScanDrop to perform water quality monitoring via the detection of an indicator coliform bacterium, Escherichia coli, in drinking water contaminated with feces. Magnetic beads conjugated with antibodies to E. coli antigen were used to selectively capture and isolate specific bacteria from water samples. The bead-captured bacteria were co-encapsulated in pico-liter droplets with fluorescently-labeled anti-E. coli antibodies, and imaged with an automated custom designed fluorescence microscope. The entire water quality diagnostic process required 8 hours from sample collection to online-accessible results compared with 2-4 days for other currently available standard detection methods.

Cloud-Enabled Microscopy and Droplet Microfluidic Platform for Specific Detection of Escherichia coli in Water

PubMed Central

Kravets, Ilia; Stawski, Nina; Hillson, Nathan J.; Yarmush, Martin L.; Marks, Robert S.; Konry, Tania

2014-01-01

We report an all-in-one platform – ScanDrop – for the rapid and specific capture, detection, and identification of bacteria in drinking water. The ScanDrop platform integrates droplet microfluidics, a portable imaging system, and cloud-based control software and data storage. The cloud-based control software and data storage enables robotic image acquisition, remote image processing, and rapid data sharing. These features form a “cloud” network for water quality monitoring. We have demonstrated the capability of ScanDrop to perform water quality monitoring via the detection of an indicator coliform bacterium, Escherichia coli, in drinking water contaminated with feces. Magnetic beads conjugated with antibodies to E. coli antigen were used to selectively capture and isolate specific bacteria from water samples. The bead-captured bacteria were co-encapsulated in pico-liter droplets with fluorescently-labeled anti-E. coli antibodies, and imaged with an automated custom designed fluorescence microscope. The entire water quality diagnostic process required 8 hours from sample collection to online-accessible results compared with 2–4 days for other currently available standard detection methods. PMID:24475107

SERS based immuno-microwell arrays for multiplexed detection of foodborne pathogenic bacteria

NASA Astrophysics Data System (ADS)

Sun, Jian; Hankus, Mikella E.; Cullum, Brian M.

2009-05-01

A novel surface enhanced Raman scattering (SERS)-based immuno-microwell array has been developed for multiplexed detection of foodborne pathogenic bacteria. The immuno-microwell array was prepared by immobilizing the optical addressable immunomagnetic beads (IMB) into the microwell array on one end of a fiber optic bundle. The IMBs, magnetic beads coated with specific antibody to specific bacteria, were used for immunomagnetic separation (IMS) of corresponding bacteria. The magnetic separation by the homemade magnetic separation system was evaluated in terms of the influences of several important parameters including the beads concentration, the sample volume and the separation time. IMS separation efficiency of the model bacteria E.coli O157:H7 was 63% in 3 minutes. The microwell array was fabricated on hydrofluoric acid etched end of a fiber optic bundle containing 30,000 fiber elements. After being coated with silver, the microwell array was used as a uniform SERS substrate with the relative standard deviation of the SERS enhancement across the microwell array < 2% and the enhancement factor as high as 2.18 x 107. The antibody modified microwell array was prepared for bacteria immobilization into the microwell array, which was characterized by a sandwich immunoassay. To demonstrate the potential of multiplexed SERS detection with the immuno-microwell array, the SERS spectra of different Raman dye labeled magnetic beads as well as mixtures were measured on the mircrowell array. In bead mixture, different beads were identified by the characteristic SERS bands of the corresponding Raman label.

A novel universal DNA labeling and amplification system for rapid microarray-based detection of 117 antibiotic resistance genes in Gram-positive bacteria.

PubMed

Strauss, Christian; Endimiani, Andrea; Perreten, Vincent

2015-01-01

A rapid and simple DNA labeling system has been developed for disposable microarrays and has been validated for the detection of 117 antibiotic resistance genes abundant in Gram-positive bacteria. The DNA was fragmented and amplified using phi-29 polymerase and random primers with linkers. Labeling and further amplification were then performed by classic PCR amplification using biotinylated primers specific for the linkers. The microarray developed by Perreten et al. (Perreten, V., Vorlet-Fawer, L., Slickers, P., Ehricht, R., Kuhnert, P., Frey, J., 2005. Microarray-based detection of 90 antibiotic resistance genes of gram-positive bacteria. J.Clin.Microbiol. 43, 2291-2302.) was improved by additional oligonucleotides. A total of 244 oligonucleotides (26 to 37 nucleotide length and with similar melting temperatures) were spotted on the microarray, including genes conferring resistance to clinically important antibiotic classes like β-lactams, macrolides, aminoglycosides, glycopeptides and tetracyclines. Each antibiotic resistance gene is represented by at least 2 oligonucleotides designed from consensus sequences of gene families. The specificity of the oligonucleotides and the quality of the amplification and labeling were verified by analysis of a collection of 65 strains belonging to 24 species. Association between genotype and phenotype was verified for 6 antibiotics using 77 Staphylococcus strains belonging to different species and revealed 95% test specificity and a 93% predictive value of a positive test. The DNA labeling and amplification is independent of the species and of the target genes and could be used for different types of microarrays. This system has also the advantage to detect several genes within one bacterium at once, like in Staphylococcus aureus strain BM3318, in which up to 15 genes were detected. This new microarray-based detection system offers a large potential for applications in clinical diagnostic, basic research, food safety and surveillance programs for antimicrobial resistance. Copyright © 2014 Elsevier B.V. All rights reserved.

Symbiotic cornucopia of the monophagous planthopper Ommatidiotus dissimilis (Fallén, 1806) (Hemiptera: Fulgoromorpha: Caliscelidae).

PubMed

Michalik, Anna; Szwedo, Jacek; Stroiński, Adam; Świerczewski, Dariusz; Szklarzewicz, Teresa

2018-03-07

In contrast to Cicadomorpha, in which numerous symbiotic bacteria have been identified and characterized, the symbionts of fulgoromorphans are poorly known. Here, we present the results of histological, ultrastructural, and molecular analyses of the symbiotic system of the planthopper Ommatidiotus dissimilis. Amplification, cloning, and sequencing of bacterial 16S RNA genes have revealed that O. dissimilis is host to five types of bacteria. Apart from bacteria Sulcia and Vidania, which are regarded as ancestral symbionts of Fulgoromorpha, three additional types of bacteria belonging to the genera Sodalis, Wolbachia, and Rickettsia have been detected. Histological and ultrastructural investigations have shown that bacteria Sulcia, Vidania, and Sodalis house separate bacteriocytes, whereas bacteria Wolbachia and Rickettsia are dispersed within various insect tissue. Additionally, bacteria belonging to the genus Vidania occupy the bacteriome localized in the lumen of the hindgut. Both molecular and microscopic analyses have revealed that all the symbionts are transovarially transmitted between generations.

Salmonella detection from chicken rinsate with surface enhanced Raman spectroscopy and RT-PCR validation

USDA-ARS?s Scientific Manuscript database

Optical detection of bacteria has been approached in recent years as a bacteria detection method that can counter time restraints of traditional plating or the high reoccurring cost of real-time polymerase chain reaction (RT-PCR). The goal of optical detection is to identify bacteria with spectral s...

Fluorescence Sensors for Early Detection of Nitrification in Drinking Water Distribution Systems - Interference Corrections and Feasibility Assessment

NASA Astrophysics Data System (ADS)

Do, T. D.; Pifer, A.; Chowdhury, Z.; Wahman, D.; Zhang, W.; Fairey, J.

2017-12-01

Detection of nitrification events in chloraminated drinking water distribution systems remains an ongoing challenge for many drinking water utilities, including Dallas Water Utilities (DWU) and the City of Houston (CoH). Each year, these utilities experience nitrification events that necessitate extensive flushing, resulting in the loss of billions of gallons of finished water. Biological techniques used to quantify the activity of nitrifying bacteria are impractical for real-time monitoring because they require significant laboratory efforts and/or lengthy incubation times. At present, DWU and CoH regularly rely on physicochemical parameters including total chlorine and monochloramine residual, and free ammonia, nitrite, and nitrate as indicators of nitrification, but these metrics lack specificity to nitrifying bacteria. To improve detection of nitrification in chloraminated drinking water distribution systems, we seek to develop a real-time fluorescence-based sensor system to detect the early onset of nitrification events by measuring the fluorescence of soluble microbial products (SMPs) specific to nitrifying bacteria. Preliminary data indicates that fluorescence-based metrics have the sensitivity to detect these SMPs in the early stages of nitrification, but several remaining challenges will be explored in this presentation. We will focus on benchtop and sensor results from ongoing batch and annular reactor experiments designed to (1) identify fluorescence wavelength pairs and data processing techniques suitable for measurement of SMPs from nitrification and (2) assess and correct potential interferences, such as those from monochloramine, pH, iron, nitrite, nitrate and humic substances. This work will serve as the basis for developing fluorescence sensor packages for full-scale testing and validation in the DWU and CoH systems. Findings from this research could be leveraged to identify nitrification events in their early stages, facilitating proactive interventions and decreasing the severity and frequency of nitrification episodes and water loss due to flushing.

Cultivation-Independent Detection of Autotrophic Hydrogen-Oxidizing Bacteria by DNA Stable-Isotope Probing ▿

PubMed Central

Pumphrey, Graham M.; Ranchou-Peyruse, Anthony; Spain, Jim C.

2011-01-01

Knallgas bacteria are a physiologically defined group that is primarily studied using cultivation-dependent techniques. Given that current cultivation techniques fail to grow most bacteria, cultivation-independent techniques that selectively detect and identify knallgas bacteria will improve our ability to study their diversity and distribution. We used stable-isotope probing (SIP) to identify knallgas bacteria in rhizosphere soil of legumes and in a microbial mat from Obsidian Pool in Yellowstone National Park. When samples were incubated in the dark, incorporation of 13CO2 was H2 dependent. SIP enabled the detection of knallgas bacteria that were not detected by cultivation, and the majority of bacteria identified in the rhizosphere soils were betaproteobacteria predominantly related to genera previously known to oxidize hydrogen. Bacteria in soil grew on hydrogen at concentrations as low as 100 ppm. A hydB homolog encoding a putative high-affinity NiFe hydrogenase was amplified from 13C-labeled DNA from both vetch and clover rhizosphere soil. The results indicate that knallgas bacteria can be detected by SIP and populations that respond to different H2 concentrations can be distinguished. The methods described here should be applicable to a variety of ecosystems and will enable the discovery of additional knallgas bacteria that are resistant to cultivation. PMID:21622787

Cultivation-independent detection of autotrophic hydrogen-oxidizing bacteria by DNA stable-isotope probing.

PubMed

Pumphrey, Graham M; Ranchou-Peyruse, Anthony; Spain, Jim C

2011-07-01

Knallgas bacteria are a physiologically defined group that is primarily studied using cultivation-dependent techniques. Given that current cultivation techniques fail to grow most bacteria, cultivation-independent techniques that selectively detect and identify knallgas bacteria will improve our ability to study their diversity and distribution. We used stable-isotope probing (SIP) to identify knallgas bacteria in rhizosphere soil of legumes and in a microbial mat from Obsidian Pool in Yellowstone National Park. When samples were incubated in the dark, incorporation of (13)CO(2) was H(2) dependent. SIP enabled the detection of knallgas bacteria that were not detected by cultivation, and the majority of bacteria identified in the rhizosphere soils were betaproteobacteria predominantly related to genera previously known to oxidize hydrogen. Bacteria in soil grew on hydrogen at concentrations as low as 100 ppm. A hydB homolog encoding a putative high-affinity NiFe hydrogenase was amplified from (13)C-labeled DNA from both vetch and clover rhizosphere soil. The results indicate that knallgas bacteria can be detected by SIP and populations that respond to different H(2) concentrations can be distinguished. The methods described here should be applicable to a variety of ecosystems and will enable the discovery of additional knallgas bacteria that are resistant to cultivation.

Microbial communities related to volatile organic compound emission in automobile air conditioning units.

PubMed

Diekmann, Nina; Burghartz, Melanie; Remus, Lars; Kaufholz, Anna-Lena; Nawrath, Thorben; Rohde, Manfred; Schulz, Stefan; Roselius, Louisa; Schaper, Jörg; Mamber, Oliver; Jahn, Dieter; Jahn, Martina

2013-10-01

During operation of mobile air conditioning (MAC) systems in automobiles, malodours can occur. We studied the microbial communities found on contaminated heat exchanger fins of 45 evaporators from car MAC systems which were operated in seven different regions of the world and identified corresponding volatile organic compounds. Collected biofilms were examined by scanning electron microscopy and fluorescent in situ hybridization. The detected bacteria were loosely attached to the metal surface. Further analyses of the bacteria using PCR-based single-strand conformation polymorphism and sequencing of isolated 16S rRNA gene fragments identified highly divergent microbial communities with multiple members of the Alphaproteobacteriales, Methylobacteria were the prevalent bacteria. In addition, Sphingomonadales, Burkholderiales, Bacillales, Alcanivorax spp. and Stenotrophomonas spp. were found among many others depending on the location the evaporators were operated. Interestingly, typical pathogenic bacteria related to air conditioning systems including Legionella spp. were not found. In order to determine the nature of the chemical compounds produced by the bacteria, the volatile organic compounds were examined by closed loop stripping analysis and identified by combined gas chromatography/mass spectrometry. Sulphur compounds, i.e. di-, tri- and multiple sulphides, acetylthiazole, aromatic compounds and diverse substituted pyrazines were detected. Mathematical clustering of the determined microbial community structures against their origin identified a European/American/Arabic cluster versus two mainly tropical Asian clusters. Interestingly, clustering of the determined volatiles against the origin of the corresponding MAC revealed a highly similar pattern. A close relationship of microbial community structure and resulting malodours to the climate and air quality at the location of MAC operation was concluded.

Colonization and Movement of Xanthomonas fragariae in Strawberry Tissues.

PubMed

Wang, Hehe; McTavish, Christine; Turechek, William W

2018-06-01

Xanthomonas fragariae causes angular leaf spot of strawberry, an important disease in strawberry growing regions worldwide. To better understand how X. fragariae multiplies and moves in strawberry plants, a green fluorescent protein (GFP)-labeled strain was constructed and used to monitor the pathogen's presence in leaf, petiole, and crown tissue with fluorescence microscopy following natural and wound inoculation in three strawberry cultivars. Taqman PCR was used to quantify bacterial densities in these same tissues regardless of the presence of GFP signal. Results showed X. fragariae colonized leaf mesophyll, the top 1 cm portion of the petiole adjacent to the leaf blade, and was occasionally found colonizing xylem vessels down to the middle of the petioles. The colonization of vascular bundles and the limited systemic movement that was observed appeared to be a passive process, of which the frequency increased with wounding and direct infiltration of bacteria into leaf veins. X. fragariae was able to directly enter petioles and colonize the space under the epidermis. Systemic movement of the bacteria into crown and other uninoculated tissues was not detected visually by GFP. However, X. fragariae was occasionally detected in these tissues by qPCR, but at quantities very near the qPCR detection limit. Petiole tissue harboring bacteria introduced either by direct entry through natural openings or wounds, or by systemic movement from infected foliar tissue, likely serves as a main source of initial inoculum in field plantings.

Optimal Conditions for the Asymmetric Polymerase Chain Reaction for Detecting Food Pathogenic Bacteria Using a Personal SPR Sensor.

PubMed

Nagai, Haruka; Tomioka, Kanji; Okumura, Shiro

2018-06-26

We have been developing quick and simple system for detecting food-poisoning bacteria using a combination of an asymmetric PCR and a portable surface plasmon resonance (SPR) sensor. The system would be suitable for point-of-care detection of food-poisoning bacteria in the field of food industry. In this study, we established a novel method for quantifying the amplified forward (F) and reverse (R) chains of Staphylococcus aureus separately by high-performance liquid chromatography (HPLC). The concentration of single-stranded DNA amplicon excessively amplified, which is crucial for the system, could be calculated as the difference between those of the F- and R-chains. For the R-chain, a correction based on the F-chain concentration in the sample was used to obtain a more accurate value, because the determination of the R-chain concentration was affected by that of the coexisting F-chain. The concentration values were also determined by fluorescence imaging for electrophoresis gels of amplicons with FITC- or Cy5-conjugated primers, and they were in good agreement with the values by the HPLC. The measured concentration of the single-strand F-chain correlated well with the value of the SPR response against the probe that was a complementary sequence of the F-chain, immobilized on the sensor chip of the SPR sensor.

Fermentation performance optimization in an ectopic fermentation system.

PubMed

Yang, Xiaotong; Geng, Bing; Zhu, Changxiong; Li, Hongna; He, Buwei; Guo, Hui

2018-07-01

Ectopic fermentation systems (EFSs) were developed for wastewater treatment. Previous studies have investigated the ability of thermophilic bacteria to improve fermentation performance in EFS. Continuing this research, we evaluated EFS performance using principle component analysis and investigated the addition of different proportions of cow dung. Viable bacteria communities were clustered and identified using BOX-AIR-based repetitive extragenic palindromic-PCR and 16S rDNA analysis. The results revealed optimal conditions for the padding were maize straw inoculated with thermophilic bacteria. Adding 20% cow dung yielded the best pH values (6.94-8.56), higher temperatures, increased wastewater absorption, improved litter quality, and greater microbial quantities. The viable bacteria groups were enriched by the addition of thermophilic consortium, and exogenous strains G21, G14, G4-1, and CR-15 were detected in fermentation process. The proportion of Bacillus species in treatment groups reached 70.37% after fermentation, demonstrating that thermophilic bacteria, especially Bacillus, have an important role in EFS, supporting previous predictions. Copyright © 2018 Elsevier Ltd. All rights reserved.

RNA-Based Methods Increase the Detection of Fecal Bacteria and Fecal Identifiers in Environmental Waters

EPA Science Inventory

We evaluated the use of qPCR RNA-based methods in the detection of fecal bacteria in environmental waters. We showed that RNA methods can increase the detection of fecal bacteria in multiple water matrices. The data suggest that this is a viable alternative for the detection of a...

ONLINE WATER MONITORING UTILIZING AN AUTOMATED MICROARRAY BIOSENSOR INSTRUMENT - PHASE I

EPA Science Inventory

Constellation Technology Corporation (Constellation) proposes the use of an integrated recovery and detection system for online water supply monitoring.  The integrated system is designed to efficiently capture and recover pathogens such as bacteria, viruses, parasites, an...

Aptasensors for rapid detection of Escherichia coli O157:H7 and Salmonella typhimurium

NASA Astrophysics Data System (ADS)

Wu, Wen-he; Li, Min; Wang, Yue; Ouyang, Hou-xian; Wang, Lin; Li, Ci-xiu; Cao, Yu-chen; Meng, Qing-he; Lu, Jian-xin

2012-11-01

Herein we reported the development of aptamer-based biosensors (aptasensors) based on label-free aptamers and gold nanoparticles (AuNPs) for detection of Escherichia coli ( E. coli) O157:H7 and Salmonella typhimurium. Target bacteria binding aptamers are adsorbed on the surface of unmodified AuNPs to capture target bacteria, and the detection was accomplished by target bacteria-induced aggregation of the aptasensor which is associated as red-to-purple color change upon high-salt conditions. By employing anti- E. coli O157:H7 aptamer and anti- S. typhimurium aptamer, we developed a convenient and rapid approach that could selectively detect bacteria without specialized instrumentation and pretreatment steps such as cell lysis. The aptasensor could detect as low as 105colony-forming units (CFU)/ml target bacteria within 20 min or less and its specificity was 100%. This novel method has a great potential application in rapid detection of bacteria in the near future.

Cultivation-Independent and Cultivation-Dependent Analysis of Microbes in the Shallow-Sea Hydrothermal System Off Kueishantao Island, Taiwan: Unmasking Heterotrophic Bacterial Diversity and Functional Capacity.

PubMed

Tang, Kai; Zhang, Yao; Lin, Dan; Han, Yu; Chen, Chen-Tung A; Wang, Deli; Lin, Yu-Shih; Sun, Jia; Zheng, Qiang; Jiao, Nianzhi

2018-01-01

Shallow-sea hydrothermal systems experience continuous fluctuations of physicochemical conditions due to seawater influx which generates variable habitats, affecting the phylogenetic composition and metabolic potential of microbial communities. Until recently, studies of submarine hydrothermal communities have focused primarily on chemolithoautotrophic organisms, however, there have been limited studies on heterotrophic bacteria. Here, fluorescence in situ hybridization, high throughput 16S rRNA gene amplicon sequencing, and functional metagenomes were used to assess microbial communities from the shallow-sea hydrothermal system off Kueishantao Island, Taiwan. The results showed that the shallow-sea hydrothermal system harbored not only autotrophic bacteria but abundant heterotrophic bacteria. The potential for marker genes sulfur oxidation and carbon fixation were detected in the metagenome datasets, suggesting a role for sulfur and carbon cycling in the shallow-sea hydrothermal system. Furthermore, the presence of diverse genes that encode transporters, glycoside hydrolases, and peptidase indicates the genetic potential for heterotrophic utilization of organic substrates. A total of 408 cultivable heterotrophic bacteria were isolated, in which the taxonomic families typically associated with oligotrophy, copiotrophy, and phototrophy were frequently found. The cultivation-independent and -dependent analyses performed herein show that Alphaproteobacteria and Gammaproteobacteria represent the dominant heterotrophs in the investigated shallow-sea hydrothermal system. Genomic and physiological characterization of a novel strain P5 obtained in this study, belonging to the genus Rhodovulum within Alphaproteobacteria, provides an example of heterotrophic bacteria with major functional capacity presented in the metagenome datasets. Collectively, in addition to autotrophic bacteria, the shallow-sea hydrothermal system also harbors many heterotrophic bacteria with versatile genetic potential to adapt to the unique environmental conditions.

Cultivation-Independent and Cultivation-Dependent Analysis of Microbes in the Shallow-Sea Hydrothermal System Off Kueishantao Island, Taiwan: Unmasking Heterotrophic Bacterial Diversity and Functional Capacity

PubMed Central

Tang, Kai; Zhang, Yao; Lin, Dan; Han, Yu; Chen, Chen-Tung A.; Wang, Deli; Lin, Yu-Shih; Sun, Jia; Zheng, Qiang; Jiao, Nianzhi

2018-01-01

Shallow-sea hydrothermal systems experience continuous fluctuations of physicochemical conditions due to seawater influx which generates variable habitats, affecting the phylogenetic composition and metabolic potential of microbial communities. Until recently, studies of submarine hydrothermal communities have focused primarily on chemolithoautotrophic organisms, however, there have been limited studies on heterotrophic bacteria. Here, fluorescence in situ hybridization, high throughput 16S rRNA gene amplicon sequencing, and functional metagenomes were used to assess microbial communities from the shallow-sea hydrothermal system off Kueishantao Island, Taiwan. The results showed that the shallow-sea hydrothermal system harbored not only autotrophic bacteria but abundant heterotrophic bacteria. The potential for marker genes sulfur oxidation and carbon fixation were detected in the metagenome datasets, suggesting a role for sulfur and carbon cycling in the shallow-sea hydrothermal system. Furthermore, the presence of diverse genes that encode transporters, glycoside hydrolases, and peptidase indicates the genetic potential for heterotrophic utilization of organic substrates. A total of 408 cultivable heterotrophic bacteria were isolated, in which the taxonomic families typically associated with oligotrophy, copiotrophy, and phototrophy were frequently found. The cultivation-independent and -dependent analyses performed herein show that Alphaproteobacteria and Gammaproteobacteria represent the dominant heterotrophs in the investigated shallow-sea hydrothermal system. Genomic and physiological characterization of a novel strain P5 obtained in this study, belonging to the genus Rhodovulum within Alphaproteobacteria, provides an example of heterotrophic bacteria with major functional capacity presented in the metagenome datasets. Collectively, in addition to autotrophic bacteria, the shallow-sea hydrothermal system also harbors many heterotrophic bacteria with versatile genetic potential to adapt to the unique environmental conditions. PMID:29527196

Ultraviolet-Absorption Spectroscopic Biofilm Monitor

NASA Technical Reports Server (NTRS)

Micheels, Ronald H.

2004-01-01

An ultraviolet-absorption spectrometer system has been developed as a prototype instrument to be used in continuous, real-time monitoring to detect the growth of biofilms. Such monitoring is desirable because biofilms are often harmful. For example, biofilms in potable-water and hydroponic systems act as both sources of pathogenic bacteria that resist biocides and as a mechanism for deterioration (including corrosion) of pipes. Biofilms formed from several types of hazardous bacteria can thrive in both plant-growth solutions and low-nutrient media like distilled water. Biofilms can also form in condensate tanks in air-conditioning systems and in industrial heat exchangers. At present, bacteria in potable-water and plant-growth systems aboard the space shuttle (and previously on the Mir space station) are monitored by culture-plate counting, which entails an incubation period of 24 to 48 hours for each sample. At present, there are no commercially available instruments for continuous monitoring of biofilms in terrestrial or spaceborne settings.

Detection and categorization of bacteria habitats using shallow linguistic analysis

PubMed Central

2015-01-01

Background Information regarding bacteria biotopes is important for several research areas including health sciences, microbiology, and food processing and preservation. One of the challenges for scientists in these domains is the huge amount of information buried in the text of electronic resources. Developing methods to automatically extract bacteria habitat relations from the text of these electronic resources is crucial for facilitating research in these areas. Methods We introduce a linguistically motivated rule-based approach for recognizing and normalizing names of bacteria habitats in biomedical text by using an ontology. Our approach is based on the shallow syntactic analysis of the text that include sentence segmentation, part-of-speech (POS) tagging, partial parsing, and lemmatization. In addition, we propose two methods for identifying bacteria habitat localization relations. The underlying assumption for the first method is that discourse changes with a new paragraph. Therefore, it operates on a paragraph-basis. The second method performs a more fine-grained analysis of the text and operates on a sentence-basis. We also develop a novel anaphora resolution method for bacteria coreferences and incorporate it with the sentence-based relation extraction approach. Results We participated in the Bacteria Biotope (BB) Task of the BioNLP Shared Task 2013. Our system (Boun) achieved the second best performance with 68% Slot Error Rate (SER) in Sub-task 1 (Entity Detection and Categorization), and ranked third with an F-score of 27% in Sub-task 2 (Localization Event Extraction). This paper reports the system that is implemented for the shared task, including the novel methods developed and the improvements obtained after the official evaluation. The extensions include the expansion of the OntoBiotope ontology using the training set for Sub-task 1, and the novel sentence-based relation extraction method incorporated with anaphora resolution for Sub-task 2. These extensions resulted in promising results for Sub-task 1 with a SER of 68%, and state-of-the-art performance for Sub-task 2 with an F-score of 53%. Conclusions Our results show that a linguistically-oriented approach based on the shallow syntactic analysis of the text is as effective as machine learning approaches for the detection and ontology-based normalization of habitat entities. Furthermore, the newly developed sentence-based relation extraction system with the anaphora resolution module significantly outperforms the paragraph-based one, as well as the other systems that participated in the BB Shared Task 2013. PMID:26201262

Sensitive detection of maltose and glucose based on dual enzyme-displayed bacteria electrochemical biosensor.

PubMed

Liu, Aihua; Lang, Qiaolin; Liang, Bo; Shi, Jianguo

2017-01-15

Glucoamylase-displayed bacteria (GA-bacteria) and glucose dehydrogenase-displayed bacteria (GDH-bacteria) were co-immobilized on multi-walled carbon nanotubes (MWNTs) modified glassy carbon electrode (GCE) to construct GA-bacteria/GDH-bacteria/MWNTs/GCE biosensor. The biosensor was developed by optimizing the loading amount and the ratio of GA-bacteria to GDH-bacteria. The as-prepared biosensor exhibited a wide dynamic range of 0.2-10mM and a low detection limit of 0.1mM maltose (S/N=3). The biosensor also had a linear response to glucose in the range of 0.1-2.0mM and a low detection limit of 0.04mM glucose (S/N=3). Interestingly, at the same concentration, glucose was 3.75-fold sensitive than that of maltose at the proposed biosensor. No interferences were observed for other possible mono- and disaccharides. The biosensor also demonstrated good long-term storage stability and repeatability. Further, using both GDH-bacteria/MWNTs/GCE biosensor and GA-bacteria/GDH-bacteria/MWNTs/GCE biosensor, glucose and maltose in real samples can be detected. Therefore, the proposed biosensor is capable of monitoring the food manufacturing and fermentation process. Copyright © 2016 Elsevier B.V. All rights reserved.

Detection of and Response to Signals Involved in Host-Microbe Interactions by Plant-Associated Bacteria

PubMed Central

Brencic, Anja; Winans, Stephen C.

2005-01-01

Diverse interactions between hosts and microbes are initiated by the detection of host-released chemical signals. Detection of these signals leads to altered patterns of gene expression that culminate in specific and adaptive changes in bacterial physiology that are required for these associations. This concept was first demonstrated for the members of the family Rhizobiaceae and was later found to apply to many other plant-associated bacteria as well as to microbes that colonize human and animal hosts. The family Rhizobiaceae includes various genera of rhizobia as well as species of Agrobacterium. Rhizobia are symbionts of legumes, which fix nitrogen within root nodules, while Agrobacterium tumefaciens is a pathogen that causes crown gall tumors on a wide variety of plants. The plant-released signals that are recognized by these bacteria are low-molecular-weight, diffusible molecules and are detected by the bacteria through specific receptor proteins. Similar phenomena are observed with other plant pathogens, including Pseudomonas syringae, Ralstonia solanacearum, and Erwinia spp., although here the signals and signal receptors are not as well defined. In some cases, nutritional conditions such as iron limitation or the lack of nitrogen sources seem to provide a significant cue. While much has been learned about the process of host detection over the past 20 years, our knowledge is far from being complete. The complex nature of the plant-microbe interactions makes it extremely challenging to gain a comprehensive picture of host detection in natural environments, and thus many signals and signal recognition systems remain to be described. PMID:15755957

Amoeba-Resisting Bacteria and Ventilator-Associated Pneumonia

PubMed Central

La Scola, Bernard; Boyadjiev, Ioanna; Greub, Gilbert; Khamis, Atieh; Martin, Claude

2003-01-01

To evaluate the role of amoeba-associated bacteria as agents of ventilator-associated pneumonia (VAP), we tested the water from an intensive care unit (ICU) every week for 6 months for such bacteria isolates; serum samples and bronchoalveolar lavage samples (BAL) were also obtained from 30 ICU patients. BAL samples were examined for amoeba-associated bacteria DNA by suicide-polymerase chain reaction, and serum samples were tested against ICU amoeba-associated bacteria. A total of 310 amoeba-associated bacteria from10 species were isolated. Twelve of 30 serum samples seroconverted to one amoeba-associated bacterium isolated in the ICU, mainly Legionella anisa and Bosea massiliensis, the most common isolates from water (p=0.021). Amoeba-associated bacteria DNA was detected in BAL samples from two patients whose samples later seroconverted. Seroconversion was significantly associated with VAP and systemic inflammatory response syndrome, especially in patients for whom no etiologic agent was found by usual microbiologic investigations. Amoeba-associated bacteria might be a cause of VAP in ICUs, especially when microbiologic investigations are negative. PMID:12890321

Bacterial detection: from microscope to smartphone.

PubMed

Gopinath, Subash C B; Tang, Thean-Hock; Chen, Yeng; Citartan, Marimuthu; Lakshmipriya, Thangavel

2014-10-15

The ubiquitous nature of bacteria enables them to survive in a wide variety of environments. Hence, the rise of various pathogenic species that are harmful to human health raises the need for the development of accurate sensing systems. Sensing systems are necessary for diagnosis and epidemiological control of pathogenic organism, especially in the food-borne pathogen and sanitary water treatment facility' bacterial populations. Bacterial sensing for the purpose of diagnosis can function in three ways: bacterial morphological visualization, specific detection of bacterial component and whole cell detection. This paper provides an overview of the currently available bacterial detection systems that ranges from microscopic observation to state-of-the-art smartphone-based detection. Copyright © 2014 Elsevier B.V. All rights reserved.

Biosensors for plant pathogen detection.

PubMed

Khater, Mohga; de la Escosura-Muñiz, Alfredo; Merkoçi, Arben

2017-07-15

Infectious plant diseases are caused by pathogenic microorganisms such as fungi, bacteria, viruses, viroids, phytoplasma and nematodes. Worldwide, plant pathogen infections are among main factors limiting crop productivity and increasing economic losses. Plant pathogen detection is important as first step to manage a plant disease in greenhouses, field conditions and at the country boarders. Current immunological techniques used to detect pathogens in plant include enzyme-linked immunosorbent assays (ELISA) and direct tissue blot immunoassays (DTBIA). DNA-based techniques such as polymerase chain reaction (PCR), real time PCR (RT-PCR) and dot blot hybridization have also been proposed for pathogen identification and detection. However these methodologies are time-consuming and require complex instruments, being not suitable for in-situ analysis. Consequently, there is strong interest for developing new biosensing systems for early detection of plant diseases with high sensitivity and specificity at the point-of-care. In this context, we revise here the recent advancement in the development of advantageous biosensing systems for plant pathogen detection based on both antibody and DNA receptors. The use of different nanomaterials such as nanochannels and metallic nanoparticles for the development of innovative and sensitive biosensing systems for the detection of pathogens (i.e. bacteria and viruses) at the point-of-care is also shown. Plastic and paper-based platforms have been used for this purpose, offering cheap and easy-to-use really integrated sensing systems for rapid on-site detection. Beside devices developed at research and development level a brief revision of commercially available kits is also included in this review. Copyright © 2016 Elsevier B.V. All rights reserved.

Bioaerosols in the Barcelona subway system.

PubMed

Triadó-Margarit, X; Veillette, M; Duchaine, C; Talbot, M; Amato, F; Minguillón, M C; Martins, V; de Miguel, E; Casamayor, E O; Moreno, T

2017-05-01

Subway systems worldwide transport more than 100 million people daily; therefore, air quality on station platforms and inside trains is an important urban air pollution issue. We examined the microbiological composition and abundance in space and time of bioaerosols collected in the Barcelona subway system during a cold period. Quantitative PCR was used to quantify total bacteria, Aspergillus fumigatus, influenza A and B, and rhinoviruses. Multitag 454 pyrosequencing of the 16S rRNA gene was used to assess bacterial community composition and biodiversity. The results showed low bioaerosol concentrations regarding the targeted microorganisms, although the bacterial bioburden was rather high (10 4 bacteria/m 3 ). Airborne bacterial communities presented a high degree of overlap among the different subway environments sampled (inside trains, platforms, and lobbies) and were dominated by a few widespread taxa, with Methylobacterium being the most abundant genus. Human-related microbiota in sequence dataset and ascribed to potentially pathogenic bacteria were found in low proportion (maximum values below 2% of sequence readings) and evenly detected. Hence, no important biological exposure marker was detected in any of the sampled environments. Overall, we found that commuters are not the main source of bioaerosols in the Barcelona subway system. © 2016 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Bacteria evade immune recognition via TLR13 and binding of their 23S rRNA by MLS antibiotics by the same mechanisms

PubMed Central

Hochrein, Hubertus; Kirschning, Carsten J.

2013-01-01

The immune system recognizes pathogens and other danger by means of pattern recognition receptors. Recently, we have demonstrated that the orphan Toll-like receptor 13 (TLR13) senses a defined sequence of the bacterial rRNA and that bacteria use specific mechanisms to evade macrolide lincosamide streptogramin (MLS) antibiotics detection via TLR13. PMID:23802068

A new multiplex real-time polymerase chain reaction assay for the diagnosis of periprosthetic joint infection.

PubMed

Kawamura, Masaki; Kobayashi, Naomi; Inaba, Yutaka; Choe, Hyonmin; Tezuka, Taro; Kubota, So; Saito, Tomoyuki

2017-11-01

A new multiplex real-time polymerase chain reaction (PCR) assay was developed to detect methicillin-resistant Staphylococcus (MRS) and to distinguish between gram-positive and gram-negative bacteria. In this study, we validated the sensitivity and specificity of this assay with periprosthetic joint infections (PJIs) and evaluated the utility of PCR for culture-negative PJI. Forty-five samples from 23 infectious PJI cases and 106 samples from 64 non-infectious control cases were analyzed by real-time PCR using a LightCycler Nano ® system. Twenty-eight clinical samples, comprising bacteria of known species isolated consecutively in the microbiological laboratory of our hospital, were used to determine the spectrum of bacterial species that could be detected using the new multiplex primers and probes. The sensitivity and specificity of the MRS- and universal-PCR assays were 92% and 99%, and 91% and 88%, respectively. Twenty-eight species of clinically isolated bacteria were detected using this method and the concordance rate for the identification of gram-positive or gram-negative organisms was 96%. Eight samples were identified as PCR-positive despite a culture-negative result. This novel multiplex real-time PCR system has acceptable sensitivity and specificity and several advantages; therefore, it has potential use for the diagnosis of PJIs, particularly in culture-negative cases.

Full-scale studies of factors related to coliform regrowth in drinking water.

PubMed

LeChevallier, M W; Welch, N J; Smith, D B

1996-07-01

An 18-month survey of 31 water systems in North America was conducted to determine the factors that contribute to the occurrence of coliform bacteria in drinking water. The survey included analysis of assimilable organic carbon (AOC), coliforms, disinfectant residuals, and operational parameters. Coliform bacteria were detected in 27.8% of the 2-week sampling periods and were associated with the following factors: filtration, temperature, disinfectant type and disinfectant level, AOC level, corrosion control, and operational characteristics. Four systems in the study that used unfiltered surface water accounted for 26.6% of the total number of bacterial samples collected but 64.3% (1,013 of 1,576) of the positive coliform samples. The occurrence of coliform bacteria was significantly higher when water temperatures were > 15 degrees C. For filtered systems that used free chlorine, 0.97% of 33,196 samples contained coliform bacteria, while 0.51% of 35,159 samples from chloraminated systems contained coliform bacteria. The average density of coliform bacteria was 35 times higher in free-chlorinated systems than in chloraminated water (0.60 CFU/100 ml for free-chlorinated water compared with 0.017 CFU/100 ml for chloraminated water). Systems that maintained dead-end free chlorine levels of < 0.2 mg/liter or monochloramine levels of < 0.5 mg/liter had substantially more coliform occurrences than systems that maintained higher disinfectant residuals. Free-chlorinated systems with AOC levels greater than 100 micrograms/liter had 82% more coliform-positive samples and 19 times higher coliform levels than free-chlorinated systems with average AOC levels less than 99 micrograms/liter. Systems that maintained a phosphate-based corrosion inhibitor and limited the amount of unlined cast iron pipe had fewer coliform bacteria. Several operational characteristics of the treatment process or the distribution system were also associated with increased rates of coliform occurrence. The study concludes that the occurrence of coliform bacteria within a distribution system is dependent upon a complex interaction of chemical, physical, operational, and engineering parameters. No one factor could account for all of the coliform occurrences, and one must consider all of the parameters described above in devising a solution to the regrowth problem.

Full-scale studies of factors related to coliform regrowth in drinking water.

PubMed Central

LeChevallier, M W; Welch, N J; Smith, D B

1996-01-01

An 18-month survey of 31 water systems in North America was conducted to determine the factors that contribute to the occurrence of coliform bacteria in drinking water. The survey included analysis of assimilable organic carbon (AOC), coliforms, disinfectant residuals, and operational parameters. Coliform bacteria were detected in 27.8% of the 2-week sampling periods and were associated with the following factors: filtration, temperature, disinfectant type and disinfectant level, AOC level, corrosion control, and operational characteristics. Four systems in the study that used unfiltered surface water accounted for 26.6% of the total number of bacterial samples collected but 64.3% (1,013 of 1,576) of the positive coliform samples. The occurrence of coliform bacteria was significantly higher when water temperatures were > 15 degrees C. For filtered systems that used free chlorine, 0.97% of 33,196 samples contained coliform bacteria, while 0.51% of 35,159 samples from chloraminated systems contained coliform bacteria. The average density of coliform bacteria was 35 times higher in free-chlorinated systems than in chloraminated water (0.60 CFU/100 ml for free-chlorinated water compared with 0.017 CFU/100 ml for chloraminated water). Systems that maintained dead-end free chlorine levels of < 0.2 mg/liter or monochloramine levels of < 0.5 mg/liter had substantially more coliform occurrences than systems that maintained higher disinfectant residuals. Free-chlorinated systems with AOC levels greater than 100 micrograms/liter had 82% more coliform-positive samples and 19 times higher coliform levels than free-chlorinated systems with average AOC levels less than 99 micrograms/liter. Systems that maintained a phosphate-based corrosion inhibitor and limited the amount of unlined cast iron pipe had fewer coliform bacteria. Several operational characteristics of the treatment process or the distribution system were also associated with increased rates of coliform occurrence. The study concludes that the occurrence of coliform bacteria within a distribution system is dependent upon a complex interaction of chemical, physical, operational, and engineering parameters. No one factor could account for all of the coliform occurrences, and one must consider all of the parameters described above in devising a solution to the regrowth problem. PMID:8779557

[Bile-resistant Gram-negative bacteria effect of different kinds of root decoction pieces].

PubMed

Deng, Yan; Wang, Ya-Ke; Han, Xiao-Yu; Wang, Ya-Qi; Jiang, Zhen-Yu; Yu, Zhi-Jun; Deng, Hai-Ying

2017-11-01

To investigate the microbial contamination in Chinese herbal decoction pieces with different functional types by studying the total aerobic microbial count (TAMC), and total yeast and mould count (TYMC) in 40 samples of 8 types of root decoction pieces; further evaluate the contamination load of bile-resistant Gram-negative bacteria, and identify the Gram-negative bacteria by using biochemical identification system for Gram-negative bacteria. Our results showed that the TAMC value was more than 1 000 CFU•g⁻¹ in 85% (34/40) samples, and was more than 100 CFU•g⁻¹ in 30% (12/40) samples; the contamination of bile-resistant Gram-negative bacteria was detected in 45% (18/40) of the samples. The bile-resistant Gram-negative bacteria load of seven batches of samples was N>1 000 MPN•g⁻¹. Sixteen bacterium strains including Serratia plymouthensis, Cedecea neteri, Escherichia vulneris, Klebsiella oxytoca, Enterobacter amnigenus, E. cloacae, E. sakazakii, Proteus penneri and E. gergoviae were obtained and identified. E. cloacae was the predominant bacterium that was isolated from Salviae Miltiorrhizae Radix et Rhizoma, while E. amnigenus, Yersinia pseudotuberculosis was the typical bacterium of Ophiopogonis Radix and Codonopsis Radix, respectively. All these suggested that the contamination of bile-resistant Gram-negative bacteria was severe for the root decoction pieces in Wuhan city. Microbial species have certain selection specificity for medicinal ingredients, so the type and limit of control bacteria for detection should be formulated according to the pollution type and quantity of bile-resistant Gram-negative bacteria. Copyright© by the Chinese Pharmaceutical Association.

Lysozyme as a recognition element for monitoring of bacterial population.

PubMed

Zheng, Laibao; Wan, Yi; Yu, Liangmin; Zhang, Dun

2016-01-01

Bacterial infections remain a significant challenge in biomedicine and environment safety. Increasing worldwide demand for point-of-care techniques and increasing concern on their safe development and use, require a simple and sensitive bioanalysis for pathogen detection. However, this goal is not yet achieved. A design for fluorescein isothiocyanate-labeled lysozyme (FITC-LYZ), which provides quantitative binding information for gram-positive bacteria, Micrococcus luteus, and detects pathogen concentration, is presented. The functional lysozyme is used not only as the pathogenic detection platform, but also as a tracking reagent for microbial population in antibacterial tests. A nonlinear relationship between the system response and the logarithm of the bacterial concentration was observed in the range of 1.2×10(2)-1.2×10(5) cfu mL(-1). The system has a potential for further applications and provides a facile and simple method for detection of pathogenic bacteria. Meanwhile, the fluorescein isothiocyanate -labeled lysozyme is also employed as the tracking agent for antibacterial dynamic assay, which show a similar dynamic curve compared with UV-vis test. Copyright © 2015 Elsevier B.V. All rights reserved.

Development of multiplex PCR for the detection of total coliform bacteria for Escherichia coli and Clostridium perfringens in drinking water.

PubMed

Tantawiwat, Suwalee; Tansuphasiri, Unchalee; Wongwit, Waranya; Wongchotigul, Varee; Kitayaporn, Dwip

2005-01-01

Multiplex PCR amplification of lacZ, uidA and plc genes was developed for the simultaneous detection of total coliform bacteria for Escherichia coli and Clostridium perfringens, in drinking water. Detection by agarose gel electrophoresis yielded a band of 876 bp for the lacZ gene of all coliform bacteria; a band of 147 bp for the uidA gene and a band of 876 bp for the lacZ gene of all strains of E. coli; a band of 280 bp for the p/c gene for all strains of C. perfringens; and a negative result for all three genes when tested with other bacteria. The detection limit was 100 pg for E. coli and C. perfringens, and 1 ng for coliform bacteria when measured with purified DNA. This assay was applied to the detection of these bacteria in spiked water samples. Spiked water samples with 0-1,000 CFU/ml of coliform bacteria and/or E. coli and/or C. perfringens were detected by this multiplex PCR after a pre-enrichment step to increase the sensitivity and to ensure that the detection was based on the presence of cultivable bacteria. The result of bacterial detection from the multiplex PCR was comparable with that of a standard plate count on selective medium (p=0.62). When using standard plate counts as a gold standard, the sensitivity for this test was 99.1% (95% CI 95.33, 99.98) and the specificity was 90.9 % (95% CI 75.67, 98.08). Multiplex PCR amplification with a pre-enrichment step was shown to be an effective, sensitive and rapid method for the simultaneous detection of these three microbiological parameters in drinking water.

9 CFR 113.25 - Culture media for detection of bacteria and fungi.

Code of Federal Regulations, 2010 CFR

2010-01-01

... 9 Animals and Animal Products 1 2010-01-01 2010-01-01 false Culture media for detection of bacteria and fungi. 113.25 Section 113.25 Animals and Animal Products ANIMAL AND PLANT HEALTH INSPECTION... STANDARD REQUIREMENTS Standard Procedures § 113.25 Culture media for detection of bacteria and fungi. (a...

9 CFR 113.25 - Culture media for detection of bacteria and fungi.

Code of Federal Regulations, 2011 CFR

2011-01-01

... 9 Animals and Animal Products 1 2011-01-01 2011-01-01 false Culture media for detection of bacteria and fungi. 113.25 Section 113.25 Animals and Animal Products ANIMAL AND PLANT HEALTH INSPECTION... STANDARD REQUIREMENTS Standard Procedures § 113.25 Culture media for detection of bacteria and fungi. (a...

PCR detection of thermophilic spore-forming bacteria involved in canned food spoilage.

PubMed

Prevost, S; Andre, S; Remize, F

2010-12-01

Thermophilic bacteria that form highly heat-resistant spores constitute an important group of spoilage bacteria of low-acid canned food. A PCR assay was developed in order to rapidly trace these bacteria. Three PCR primer pairs were designed from rRNA gene sequences. These primers were evaluated for the specificity and the sensitivity of detection. Two primer pairs allowed detection at the species level of Geobacillus stearothermophilus and Moorella thermoacetica/thermoautrophica. The other pair allowed group-specific detection of anaerobic thermophilic bacteria of the genera Thermoanaerobacterium, Thermoanaerobacter, Caldanerobium and Caldanaerobacter. After a single enrichment step, these PCR assays allowed the detection of 28 thermophiles from 34 cans of spoiled low-acid food. In addition, 13 ingredients were screened for the presence of these bacteria. This PCR assay serves as a detection method for strains able to spoil low-acid canned food treated at 55°C. It will lead to better reactivity in the canning industry. Raw materials and ingredients might be qualified not only for quantitative spore contamination, but also for qualitative contamination by highly heat-resistant spores.

Quantitative and simultaneous detection of four foodborne bacterial pathogens with a multi-channel SPR sensor.

PubMed

Taylor, Allen D; Ladd, Jon; Yu, Qiuming; Chen, Shengfu; Homola, Jirí; Jiang, Shaoyi

2006-12-15

We report the quantitative and simultaneous detection of four species of bacteria, Escherichia coli O157:H7, Salmonella choleraesuis serotype typhimurium, Listeria monocytogenes, and Campylobacter jejuni, using an eight-channel surface plasmon resonance (SPR) sensor based on wavelength division multiplexing. Detection curves showing SPR response versus analyte concentration were established for each species of bacteria in buffer at pH 7.4, apple juice at native pH 3.7, and apple juice at an adjusted pH of 7.4, as well as for a mixture containing all four species of bacteria in buffer. Control experiments were performed to show the non-fouling characteristics of the sensor surface as well as the specificity of the amplification antibodies used in this study. The limit of detection (LOD) for each of the four species of bacteria in the tested matrices ranges from 3.4 x 10(3) to 1.2 x 10(5) cfu/ml. Detection curves in buffer of an individual species of bacteria in a mixture of all four species of bacteria correlated well with detection curves of the individual species of bacteria alone. SPR responses were higher for bacteria in apple juice at pH 7.4 than in apple juice at pH 3.7. This difference in sensor response could be partly attributed to the pH dependence of antibody-antigen binding.

Microfluidic cantilever detects bacteria and measures their susceptibility to antibiotics in small confined volumes

NASA Astrophysics Data System (ADS)

Etayash, Hashem; Khan, M. F.; Kaur, Kamaljit; Thundat, Thomas

2016-10-01

In the fight against drug-resistant bacteria, accurate and high-throughput detection is essential. Here, a bimaterial microcantilever with an embedded microfluidic channel with internal surfaces chemically or physically functionalized with receptors selectively captures the bacteria passing through the channel. Bacterial adsorption inside the cantilever results in changes in the resonance frequency (mass) and cantilever deflection (adsorption stress). The excitation of trapped bacteria using infrared radiation (IR) causes the cantilever to deflect in proportion to the infrared absorption of the bacteria, providing a nanomechanical infrared spectrum for selective identification. We demonstrate the in situ detection and discrimination of Listeria monocytogenes at a concentration of single cell per μl. Trapped Escherichia coli in the microchannel shows a distinct nanomechanical response when exposed to antibiotics. This approach, which combines enrichment with three different modes of detection, can serve as a platform for the development of a portable, high-throughput device for use in the real-time detection of bacteria and their response to antibiotics.

Brief ultrasonication improves detection of biofilm-formative bacteria around a metal implant.

PubMed

Kobayashi, Naomi; Bauer, Thomas W; Tuohy, Marion J; Fujishiro, Takaaki; Procop, Gary W

2007-04-01

Biofilms are complex microenvironments produced by microorganisms on surfaces. Ultrasonication disrupts biofilms and may make the microorganism or its DNA available for detection. We determined whether ultrasonication could affect our ability to detect bacteria adherent to a metal substrate. A biofilm-formative Staphylococcus aureus strain was used for an in vitro implant infection model (biofilm-formative condition). We used quantitative culture and real time-polymerase chain reaction to determine the influence of different durations of ultrasound on bacterial adherence and viability. Sonication for 1 minute increased the yield of bacteria. Sonication longer than 5 minutes led to fewer bacterial colonies by conventional culture but not by polymerase chain reaction. This suggests short periods of sonication help release bacteria from the metal substrate by disrupting the biofilm, but longer periods of sonication lyse bacteria prohibiting their detection in microbiologic cultures. A relatively short duration of sonication may be desirable for maximizing detection of biofilm-formative bacteria around implants by culture or polymerase chain reaction.

Several methods for concentrating bacteria in fluid samples

NASA Technical Reports Server (NTRS)

Thomas, R. R.

1976-01-01

The sensitivities of the firefly luciferase - ATP flow system and luminol flow system were established as 300,000 E. coli per milliliter and 10,000 E. coli per milliliter respectively. To achieve the detection limit of 1,000 bacteria per milliliter previously established, a method of concentrating microorganisms using a sartorius membrane filter system is investigated. Catalase in 50% ethanol is found to be a stable luminol standard and can be used up to 24 hours with only a 10% loss of activity. The luminol reagent is also stable over a 24 hour period. A method of preparing relatively inexpensive luciferase from desiccated firefly tails is developed.

Surface immuno-functionalisation for the capture and detection of Vibrio species in the marine environment: a new management tool for industrial facilities.

PubMed

Laczka, Olivier F; Labbate, Maurizio; Seymour, Justin R; Bourne, David G; Fielder, Stewart S; Doblin, Martina A

2014-01-01

Bacteria from the genus Vibrio are a common and environmentally important group of bacteria within coastal environments and include species pathogenic to aquaculture organisms. Their distribution and abundance are linked to specific environmental parameters, including temperature, salinity and nutrient enrichment. Accurate and efficient detection of Vibrios in environmental samples provides a potential important indicator of overall ecosystem health while also allowing rapid management responses for species pathogenic to humans or species implicated in disease of economically important aquacultured fish and invertebrates. In this study, we developed a surface immuno-functionalisation protocol, based on an avidin-biotin type covalent binding strategy, allowing specific sandwich-type detection of bacteria from the Vibrio genus. The assay was optimized on 12 diverse Vibrio strains, including species that have implications for aquaculture industries, reaching detection limits between 7×10(3) to 3×10(4) cells mL(-1). Current techniques for the detection of total Vibrios rely on laborious or inefficient analyses resulting in delayed management decisions. This work represents a novel approach for a rapid, accurate, sensitive and robust tool for quantifying Vibrios directly in industrial systems and in the environment, thereby facilitating rapid management responses.

Linking fecal bacteria in rivers to landscape, geochemical, and hydrologic factors and sources at the basin scale

PubMed Central

Verhougstraete, Marc P.; Martin, Sherry L.; Kendall, Anthony D.; Hyndman, David W.; Rose, Joan B.

2015-01-01

Linking fecal indicator bacteria concentrations in large mixed-use watersheds back to diffuse human sources, such as septic systems, has met limited success. In this study, 64 rivers that drain 84% of Michigan’s Lower Peninsula were sampled under baseflow conditions for Escherichia coli, Bacteroides thetaiotaomicron (a human source-tracking marker), landscape characteristics, and geochemical and hydrologic variables. E. coli and B. thetaiotaomicron were routinely detected in sampled rivers and an E. coli reference level was defined (1.4 log10 most probable number⋅100 mL−1). Using classification and regression tree analysis and demographic estimates of wastewater treatments per watershed, septic systems seem to be the primary driver of fecal bacteria levels. In particular, watersheds with more than 1,621 septic systems exhibited significantly higher concentrations of B. thetaiotaomicron. This information is vital for evaluating water quality and health implications, determining the impacts of septic systems on watersheds, and improving management decisions for locating, constructing, and maintaining on-site wastewater treatment systems. PMID:26240328

Investigation of N-acyl homoserine lactone (AHL) molecule production in Gram-negative bacteria isolated from cooling tower water and biofilm samples.

PubMed

Haslan, Ezgi; Kimiran-Erdem, Ayten

2013-09-01

In this study, 99 Gram-negative rod bacteria were isolated from cooling tower water, and biofilm samples were examined for cell-to-cell signaling systems, N-acyl homoserine lactone (AHL) signal molecule types, and biofilm formation capacity. Four of 39 (10 %) strains isolated from water samples and 14 of 60 (23 %) strains isolated from biofilm samples were found to be producing a variety of AHL signal molecules. It was determined that the AHL signal molecule production ability and the biofilm formation capacity of sessile bacteria is higher than planktonic bacteria, and there was a statistically significant difference between the AHL signal molecule production of these two groups (p < 0.05). In addition, it was found that bacteria belonging to the same species isolated from cooling tower water and biofilm samples produced different types of AHL signal molecules and that there were different types of AHL signal molecules in an AHL extract of bacteria. In the present study, it was observed that different isolates of the same strains did not produce the same AHLs or did not produce AHL molecules, and bacteria known as AHL producers did not produce AHL. These findings suggest that detection of signal molecules in bacteria isolated from cooling towers may contribute to prevention of biofilm formation, elimination of communication among bacteria in water systems, and blockage of quorum-sensing controlled virulence of these bacteria.

Improvement in the detection rate of diarrhoeagenic bacteria in human stool specimens by a rapid real-time PCR assay.

PubMed

Iijima, Yoshio; Asako, Nahoko T; Aihara, Masanori; Hayashi, Kozaburo

2004-07-01

A rapid laboratory system has been developed and evaluated that can simultaneously identify major diarrhoeagenic bacteria, including Salmonella enterica, Vibrio parahaemolyticus, Campylobacter jejuni and Shiga toxin-producing Escherichia coli, in stool specimens by real-time PCR. Specific identification was achieved by using selective TaqMan probes, detecting two targets in each pathogen. A positive result was scored only when both targets of a pathogen were amplified and the difference between threshold cycles for detection was less than five. Diagnosis of enteric bacterial infections using this highly sensitive method, including DNA extraction and real-time PCR, requires only 3 h. Forty stool specimens related to suspected food poisoning outbreaks were analysed: 16 (40%) of these samples were found to be positive for diarrhoeagenic bacteria using a conventional culture method; 28 (70%) were positive using the real-time PCR assay. Of the 12 PCR-positive but culture-negative cases, 11 patients had consumed pathogen-contaminated or high-risk food. Analysis of faecal samples from 105 outpatients who complained of diarrhoea and/or abdominal pain identified 19 (18%) patients as being positive for diarrhoeagenic bacteria using the culture method. An additional six (6%) patients were found to be positive by PCR analysis.

Microbiological diversity and prevalence of spoilage and pathogenic bacteria in commercial fermented alcoholic beverages (beer, fruit wine, refined rice wine, and yakju).

PubMed

Jeon, Se Hui; Kim, Nam Hee; Shim, Moon Bo; Jeon, Young Wook; Ahn, Ji Hye; Lee, Soon Ho; Hwang, In Gyun; Rhee, Min Suk

2015-04-01

The present study examined 469 commercially available fermented alcoholic beverages (FABs), including beer (draft, microbrewed, and pasteurized), fruit wine (grape and others), refined rice wine, and yakju (raw and pasteurized). Samples were screened for Escherichia coli and eight foodborne pathogens (Bacillus cereus, Campylobacter jejuni, Clostridium perfringens, Escherichia coli O157:H7, Listeria monocytogenes, Salmonella spp., Staphylococcus aureus, and Yersinia enterocolitica), and the aerobic plate count, lactic acid bacteria, acetic acid bacteria, fungi, and total coliforms were also enumerated. Microbrewed beer contained the highest number of microorganisms (average aerobic plate count, 3.5; lactic acid bacteria, 2.1; acetic acid bacteria, 2.0; and fungi, 3.6 log CFU/ml), followed by draft beer and yakju (P < 0.05), whereas the other FABs contained , 25 CFU/25 ml microorganisms. Unexpectedly, neither microbial diversity nor microbial count correlated with the alcohol content (4.7 to 14.1%) or pH (3.4 to 4.2) of the product. Despite the harsh conditions, coliforms (detected in 23.8% of microbrewed beer samples) and B. cereus (detected in all FABs) were present in some products. B. cereus was detected most frequently in microbrewed beer (54.8% of samples) and nonpasteurized yakju (50.0%), followed by pasteurized yakju (28.8%), refined rice wine (25.0%), other fruit wines (12.3%), grape wine (8.6%), draft beer (5.6%), and pasteurized beer (2.2%) (P < 0.05). The finding that spore-forming B. cereus and coliform bacteria can survive the harsh conditions present in alcoholic beverages should be taken into account (alongside traditional quality indicators such as the presence of lactic acid-producing bacteria, acetic acid-producing bacteria, or both) when developing manufacturing systems and methods to prolong the shelf life of high-quality FAB products. New strategic quality management plans for various FABs are needed.

Two techniques for eliminating luminol interference material and flow system configurations for luminol and firefly luciferase systems

NASA Technical Reports Server (NTRS)

Thomas, R. R.

1976-01-01

Two methods for eliminating luminol interference materials are described. One method eliminates interference from organic material by pre-reacting a sample with dilute hydrogen peroxide. The reaction rate resolution method for eliminating inorganic forms of interference is also described. The combination of the two methods makes the luminol system more specific for bacteria. Flow system designs for both the firefly luciferase and luminol bacteria detection systems are described. The firefly luciferase flow system incorporating nitric acid extraction and optimal dilutions has a functional sensitivity of 3 x 100,000 E. coli/ml. The luminol flow system incorporates the hydrogen peroxide pretreatment and the reaction rate resolution techniques for eliminating interference. The functional sensitivity of the luminol flow system is 1 x 10,000 E. coli/ml.

Detection of proteins and bacteria using an array of feedback capacitance sensors.

PubMed

Mehta, Manav; Hanumanthaiah, Chandra Sekar; Betala, Pravin Ajitkumar; Zhang, Hong; Roh, SaeWeon; Buttner, William; Penrose, William R; Stetter, Joseph R; Pérez-Luna, Victor H

2007-12-15

An integrated array of micron-dimension capacitors, originally developed for biometric applications (fingerprint identification), was engineered for detection of biological agents such as proteins and bacteria. This device consists of an array of 93,184 (256 x 364) individual capacitor-based sensing elements located underneath a thin (0.8 microm) layer of glass. This glass layer can be functionalized with organosilane-based monolayers to provide groups amenable for the immobilization of bioreceptors such as antibodies, enzymes, peptides, aptamers, and nucleotides. Upon functionalization with antibodies and in conjunction with signal amplification schemes that result in perturbation of the dielectric constant around the captured antigens, this system can be used as a detector of biological agents. Two signal amplification schemes were tested in this work: one consisted of 4 microm diameter latex immunobeads and a second one was based on colloidal gold catalyzed reduction of silver. These signal amplification approaches were demonstrated and show that this system is capable of specific detection of bacteria (Escherichia coli) and proteins (ovalbumin). The present work shows proof-of-principle demonstration that a simple fingerprint detector based on feedback capacitance measurements can be implemented as a biosensor. The approach presented could be easily expanded to simultaneously test for a large number of analytes and multiple samples given that this device has a large number of detectors. The device and required instrumentation is highly portable and does not require expensive and bulky instrumentation because it relies purely on electronic detection.

Natural soil reservoirs for human pathogenic and fecal indicator bacteria

USGS Publications Warehouse

Boschiroli, Maria L; Falkinham, Joseph; Favre-Bonte, Sabine; Nazaret, Sylvie; Piveteau, Pascal; Sadowsky, Michael J.; Byappanahalli, Muruleedhara; Delaquis, Pascal; Hartmann, Alain

2016-01-01

Soils receive inputs of human pathogenic and indicator bacteria through land application of animal manures or sewage sludge, and inputs by wildlife. Soil is an extremely heterogeneous substrate and contains meso- and macrofauna that may be reservoirs for bacteria of human health concern. The ability to detect and quantify bacteria of human health concern is important in risk assessments and in evaluating the efficacy of agricultural soil management practices that are protective of crop quality and protective of adjacent water resources. The present chapter describes the distribution of selected Gram-positive and Gram-negative bacteria in soils. Methods for detecting and quantifying soilborne bacteria including extraction, enrichment using immunomagnetic capture, culturing, molecular detection and deep sequencing of metagenomic DNA to detect pathogens are overviewed. Methods for strain phenotypic and genotypic characterization are presented, as well as how comparison with clinical isolates can inform the potential for human health risk.

Oral bacteria influenced by the functional status of the elderly people and the type and quality of facilities for the bedridden.

PubMed

Tada, A; Watanabe, T; Yokoe, H; Hanada, N; Tanzawa, H

2002-01-01

To analyse the relationship between oral bacteria and the health and functional status of the elderly. The bacteria species in the oral cavity of the elderly were examined. It was found that the bedridden subjects staying at two hospitals for long-term (HOBR) showed significantly lower detection rates of commensal bacteria species and significantly higher detection rates of Pseudomonas aeruginosa, of methicillin-resistant Staphylococcus aureus (MRSA) and of coagulase(-) Staph. aureus than those living independently (the independent). In addition, the detection rate of Haemophilus parainfluenzae in NUBR was discovered to be higher than that found in the independent. In HOBR, the detection rate of Ps. aeruginosa was significantly higher among in-patients who required continual care than those in need of partial care, while the detection rate of MRSA was significantly higher among in-patients with low serum albumin than those with normal serum albumin. Oral bacteria examination analysis proved that the risks of infection of some pathogenic bacteria species were correlated with functional status, physical function and nutritional state. Our study suggests that the oral bacteria, especially pathogenic bacteria were influenced by the functional status of the elderly and the type and quality of the facilities for the bedridden, physical function and nutritional state.

Rapid quantification of live/dead lactic acid bacteria in probiotic products using high-sensitivity flow cytometry

NASA Astrophysics Data System (ADS)

He, Shengbin; Hong, Xinyi; Huang, Tianxun; Zhang, Wenqiang; Zhou, Yingxing; Wu, Lina; Yan, Xiaomei

2017-06-01

A laboratory-built high-sensitivity flow cytometer (HSFCM) was employed for the rapid and accurate detection of lactic acid bacteria (LAB) and their viability in probiotic products. LAB were stained with both the cell membrane-permeable SYTO 9 green-fluorescent nucleic acid stain and the red-fluorescent nucleic acid stain, propidium iodide, which penetrates only bacteria with compromised membranes. The side scatter and dual-color fluorescence signals of single bacteria were detected simultaneously by the HSFCM. Ultra-high temperature processing milk and skim milk spiked with Lactobacillus casei were used as the model systems for the optimization of sample pretreatment and staining. The viable LAB counts measured by the HSFCM were in good agreement with those of the plate count method, and the measured ratios between the live and dead LAB matched well with the theoretical ratios. The established method was successfully applied to the rapid quantification of live/dead LAB in yogurts and fermented milk beverages of different brands. Moreover, the concentration and viability status of LAB in ambient yogurt, a relatively new yet popular milk product in China, are also reported.

Predominant bacteria in an activated sludge reactor for the degradation of cutting fluids

DOE Office of Scientific and Technical Information (OSTI.GOV)

Baker, C.A.; Claus, G.W.; Taylor, P.A.

1983-01-01

For the first time, an activated sludge reactor, established for the degradation of cutting fluids, was examined for predominant bacteria. In addition, both total and viable numbers of bacteria in the reactor were determined so that the percentage of each predominant type in the total reactor population could be determined. Three samples were studied, and a total of 15 genera were detected. In each sample, the genus Pseudomonas and the genus Microcyclus were present in high numbers. Three other genera, Acinetobacter, Alcaligenes, and Corynebacterium, were also found in every sample but in lower numbers. In one sample, numerous appendage bacteriamore » were present, and one of these, the genus Seliberia, was the most predominant organism in that sample. However, in the other two samples no appendage bacteria were detected. Six genera were found in this reactor which have not been previously reported in either cutting fluids in use or in other activated sludge systems. These genera were Aeromonas, Hyphomonas, Listeria, Microcyclus, Moraxella, and Spirosoma. None of the predominant bacterial belonged to groups of strict pathogens. 22 references, 6 figures, 3 tables.« less

Detection of phenols using engineered bacteria

DOEpatents

Wise, Arlene A.; Kuske, Cheryl R.; Terwilliger, Thomas C.

2007-12-04

Detection of phenols using engineered bacteria. A biosensor can be created by placing a reporter gene under control of an inducible promoter. The reporter gene produces a signal when a cognate transcriptional activator senses the inducing chemical. Creation of bacterial biosensors is currently restricted by limited knowledge of the genetic systems of bacteria that catabolize xenobiotics. By using mutagenic PCR to change the chemical specificity of the Pseudomonas species CF600 DmpR protein, the potential for engineering novel biosensors for detection of phenols has been demonstrated. DmpR, a well-characterized transcriptional activator of the P. CF600's dmp operon mediates growth on simple phenols. Transcription from Po, the promoter heading the dmp operon, is activated when the sensor domain of DmpR interacts with phenol and mono-substituted phenols. By altering the sensor domain of the DmpR, a group of DmpR derivatives that activate transcription of a Po-lacZ fusion in response to eight of the EPA's eleven priority pollutant phenols has been created. The assays and the sensor domain mutations that alter the chemical specificity of DmpR is described.

Detection of phenols using engineered bacteria

DOEpatents

Wise, Arlene A.; Kuske, Cheryl R.; Terwilliger, Thomas C.

2004-08-10

Detection of phenols using engineered bacteria. A biosensor can be created by placing a reporter gene under control of an inducible promoter. The reporter gene produces a signal when a cognate transcriptional activator senses the inducing chemical. Creation of bacterial biosensors is currently restricted by limited knowledge of the genetic systems of bacteria that catabolize xenobiotics. By using mutagenic PCR to change the chemical specificity of the Pseudomonas species CF600 DmpR protein, the potential for engineering novel biosensors for detection of phenols has been demonstrated. DmpR, a well-characterized transcriptional activator of the P. CF600's dmp operon mediates growth on simple phenols. Transcription from Po, the promoter heading the dmp operon, is activated when the sensor domain of DmpR interacts with phenol and mono-substituted phenols. By altering the sensor domain of the DmpR, a group of DmpR derivatives that activate transcription of a Po-lacZ fusion in response to eight of the EPA's eleven priority pollutant phenols has been created. The assays and the sensor domain mutations that alter the chemical specificity of DmpR is described.

Detection of periodontal bacteria in thrombi of patients with acute myocardial infarction by polymerase chain reaction.

PubMed

Ohki, Takahiro; Itabashi, Yuji; Kohno, Takashi; Yoshizawa, Akihiro; Nishikubo, Shuichi; Watanabe, Shinya; Yamane, Genyuki; Ishihara, Kazuyuki

2012-02-01

Numerous reports have demonstrated that periodontal bacteria are present in plaques from atherosclerotic arteries. Although periodontitis has recently been recognized as a risk factor for coronary artery disease, the direct relationship between periodontal bacteria and coronary artery disease has not yet been clarified. It has been suggested that these bacteria might contribute to inflammation and plaque instability. We assumed that if periodontal bacteria induce inflammation of plaque, the bacteria would be released into the bloodstream when vulnerable plaque ruptures. To determine whether periodontal bacteria are present in thrombi at the site of acute myocardial infarction, we tried to detect periodontal bacteria in thrombi of patients with acute myocardial infarction by polymerase chain reaction (PCR). We studied 81 consecutive adults with ST-segment elevation acute myocardial infarction who underwent primary percutaneous coronary intervention (PCI). All patients underwent removal of thrombus with aspiration catheters at the beginning of percutaneous coronary intervention, and a small sample of thrombus was obtained for PCR. The detection rates of periodontal bacteria by PCR were 19.7% for Aggregatibacter actinomycetemcomitans, 3.4% for Porphyromonas gingivalis, and 2.3% for Treponema denticola. Three species of periodontal bacteria were detected in the thrombi of patients with acute myocardial infarction. This raises the possibility that such bacteria are latently present in plaque and also suggests that these bacteria might have a role in plaque inflammation and instability. Copyright © 2012 Mosby, Inc. All rights reserved.

Multiple Strategy Bio-Detection Sensor Platforms Made From Carbon and Polymer Materials

DTIC Science & Technology

2006-08-10

binding of antigens. Further investigations were conducted on the antibody-antigen detection scheme using known test antigens (i.e. bacteria ). While...this method worked in preliminary studies, the use of antibodies was determined not to be feasible for detecting many different types of bacteria in...solution. Also, as the number of bacteria in a solution increased, it became necessary to wash the detection surface more extensively to favor specific

Impedimetric detection of pathogenic Gram-positive bacteria using an antimicrobial peptide from class IIa bacteriocins.

PubMed

Etayash, Hashem; Jiang, Keren; Thundat, Thomas; Kaur, Kamaljit

2014-02-04

Real-time, label-free detection of Gram-positive bacteria with high selectivity and sensitivity is demonstrated using an interdigitated impedimetric array functionalized with naturally produced antimicrobial peptide from class IIa bacteriocins. The antimicrobial peptide, leucocin A, was chemically synthesized and covalently immobilized on interdigitated gold microelectrodes via the interaction between the C-terminal carboxylic acid of the peptide and free amines of a preattached thiolated linker. Exposing the peptide sensor to various concentrations of Gram-positive bacteria generated reproducible impedance spectra that detected peptide-bacteria interactions at a concentration of 1 cell/μL. The peptide sensor also selectively detected Listeria monocytogenes from other Gram-positive strains at a concentration of 10(3) cfu mL(-1). The study highlights that short peptide ligands from bacteriocin class offer high selectivity in bacterial detection and can be used in developing a robust, portable biosensor device to efficiently detect pathogenic Gram-positive bacteria in food samples.

Near real time, accurate, and sensitive microbiological safety monitoring using an all-fibre spectroscopic fluorescence system

NASA Astrophysics Data System (ADS)

Vanholsbeeck, F.; Swift, S.; Cheng, M.; Bogomolny, E.

2013-11-01

Enumeration of microorganisms is an essential microbiological task for many industrial sectors and research fields. Various tests for detection and counting of microorganisms are used today. However most of the current methods to enumerate bacteria require either long incubation time for limited accuracy, or use complicated protocols along with bulky equipment. We have developed an accurate, all-fibre spectroscopic system to measure fluorescence signal in-situ. In this paper, we examine the potential of this setup for near real time bacteria enumeration in aquatic environment. The concept is based on a well-known phenomenon that the fluorescence quantum yields of some nucleic acid stains significantly increase upon binding with nucleic acids of microorganisms. In addition we have used GFP labeled organisms. The fluorescence signal increase can be correlated to the amount of nucleic acid present in the sample. In addition we have used GFP labeled organisms. Our results show that we are able to detect a wide range of bacteria concentrations without dilution or filtration (1-108 CFU/ml) using different optical probes we designed. This high sensitivity is due to efficient light delivery with an appropriate collection volume and in situ fluorescence detection as well as the use of a sensitive CCD spectrometer. By monitoring the laser power, we can account for laser fluctuations while measuring the fluorescence signal which improves as well the system accuracy. A synchronized laser shutter allows us to achieve a high SNR with minimal integration time, thereby reducing the photobleaching effect. In summary, we conclude that our optical setup may offer a robust method for near real time bacterial detection in aquatic environment.

Occurrence of Pepper Mild Mottle Virus (PMMoV) in Groundwater from a Karst Aquifer System in the Yucatan Peninsula, Mexico.

PubMed

Rosiles-González, Gabriela; Ávila-Torres, Gerardo; Moreno-Valenzuela, Oscar A; Acosta-González, Gilberto; Leal-Bautista, Rosa María; Grimaldo-Hernández, Cinthya D; Brown, Judith K; Chaidez-Quiroz, Cristóbal; Betancourt, Walter Q; Gerba, Charles P; Hernández-Zepeda, Cecilia

2017-12-01

The Yucatan Peninsula of Mexico hosts a karst aquifer system that is the only source of freshwater for the area; however, it is vulnerable to human-mediated contamination. Pepper mild mottle virus (PMMoV) is one of the most abundant RNA viruses associated with human feces, making it a viable indicator for tracking fecal pollution in aquatic environments, including groundwater. In this study, groundwater samples collected from a karst aquifer from fresh and brackish water locations were analyzed for fecal indicator bacteria, somatic and male F+ specific coliphages, and PMMoV during the rainy and dry seasons. Total coliform bacteria were detected at all sites, whereas Escherichia coli were found at relatively low levels <40 MPN/100 ml. The highest average concentrations of somatic and male F+ specific coliphages were 920 and 330 plaque forming units per 100 ml, respectively, detected in freshwater during the rainy season. PMMoV RNA was detected in 85% of the samples with gene sequences sharing 99-100% of nucleotide identity with PMMoV sequences available in GenBank. Quantification of PMMoV genome copies (GC) by quantitative real-time PCR indicated concentrations ranging from 1.7 × 10 1 to 1.0 × 10 4 GC/L, with the highest number of GC detected during the rainy season. No significant correlation was observed between PMMoV occurrence by season or water type (p > 0.05). Physicochemical and indicator bacteria were not correlated with PMMoV concentrations. The abundance and prevalence of PMMoV in the karst aquifer may reflect its environmental persistence and its potential as a fecal indicator in this karst aquifer system.

Comprehensive evaluation of SCFA production in the intestinal bacteria regulated by berberine using gas-chromatography combined with polymerase chain reaction.

PubMed

Wang, Lu-Lu; Guo, Hui-Hui; Huang, Shuai; Feng, Chen-Lin; Han, Yan-Xing; Jiang, Jian-Dong

2017-07-01

Short-chain fatty acids (SCFAs) of intestine microbial have caught accumulating attention for their beneficial effects on human health. Botanic compounds with low bioavailability such as berberine (BBR) and resveratrol might interact with intestinal microbial ecosystem and promote gut bacteria to produce SCFA, which contribute to their biological effects. In the present study, a comprehensive assay system was built to detect SCFAs production in intestinal bacteria, in which stringent anaerobic culture was applied for in vitro bacterial fermentation, followed by direct-injection GC detection (chemical detection) in combination with real time polymerase chain reaction (RT-PCR, biological detection). BBR was used as positive reference. The direct injection GC method was calibrated and successfully applied to analyze the concentration of SCFAs in gut microbiota and BBR was proved to be effective in the dose- and time-dependent up-regulation of SCFAs production. As compared to the saline group, the concentration of acetic acid, propionate acid and butyric acid (the main SCFAs in gut microbiota) were increased by 17.7%, 11.1% and 30.5%, respectively, after incubating intestinal bacteria with 20μg/mL BBR for 24h. The increase reached to 34.9%, 22.4% and 51.6%, respectively when the BBR was 50μg/mL. Additionally, consensus-degenerate hybrid oligonucleotide primers (CODEHOPs) were designed for the detection of acetate kinase (ACK), Methylmalonyl-CoA decarboxylase (MMD) and butyryl-CoA: acetate-CoA transferase (BUT), as they are the key enzymes in the synthetic pathway for acetic acid, propionate acid and butyric acid, respectively. After 24hr's incubation, BBR was shown to promote the gene expression of ACK, MMD and BUT significantly (86.5%, 27.2% and 60.4%, respectively, with 20μg/mL BBR; 130.2%, 84.2% and 98.4%, respectively, with 50μg/mL BBR), showing a solid biological support for the chemical detection. This comprehensive assay system might be useful in identifying SCFAs promoting agents with information on their mechanism. Copyright © 2017 Elsevier B.V. All rights reserved.

Validating tyrosinase homologue MelA as a photoacoustic reporter gene for imaging Escherichia coli

NASA Astrophysics Data System (ADS)

Paproski, Robert J.; Li, Yan; Barber, Quinn; Lewis, John D.; Campbell, Robert; Zemp, Roger

2015-03-01

Antibiotic drug resistance is a major worldwide issue. Development of new therapies against pathogenic bacteria requires appropriate research tools for replicating and characterizing infections. Previously fluorescence and bioluminescence modalities have been used to image infectious burden in animal models but scattering significantly limits imaging depth and resolution. We hypothesize that photoacoustic imaging, which has improved depth-toresolution ratio, could be useful for visualizing MelA-expressing bacteria since MelA is a bacterial tyrosinase homologue involved in melanin production. Using an inducible expression system, E. coli expressing MelA were visibly black in liquid culture. Phosphate buffered saline (PBS), MelA-expressing bacteria (at different dilutions in PBS), and chicken embryo blood were injected in plastic tubes which were imaged using a VisualSonics Vevo LAZR system. Photoacoustic imaging at 6 different wavelengths (680, 700, 750, 800, 850 and 900nm) enabled spectral de-mixing to distinguish melanin signals from blood. The signal to noise ratio of 9x diluted MelA bacteria was 55, suggesting that ~20 bacteria cells could be detected with our system. When MelA bacteria were injected as a 100 μL bolus into a chicken embryo, photoacoustic signals from deoxy- and oxy- hemoglobin as well as MelA-expressing bacteria could be separated and overlaid on an ultrasound image, allowing visualization of the bacterial location. Photoacoustic imaging may be a useful tool for visualizing bacterial infections and further work incorporating photoacoustic reporters into infectious bacterial strains is warranted.

Trace detection of specific viable bacteria using tetracysteine-tagged bacteriophages.

PubMed

Wu, Lina; Luan, Tian; Yang, Xiaoting; Wang, Shuo; Zheng, Yan; Huang, Tianxun; Zhu, Shaobin; Yan, Xiaomei

2014-01-07

Advanced methods are urgently needed to determine the identity and viability of trace amounts of pathogenic bacteria in a short time. Existing approaches either fall short in the accurate assessment of microbial viability or lack specificity in bacterial identification. Bacteriophages (or phages for short) are viruses that exclusively infect bacterial host cells with high specificity. As phages infect and replicate only in living bacterial hosts, here we exploit the strategy of using tetracysteine (TC)-tagged phage in combination with biarsenical dye to the discriminative detection of viable target bacteria from dead target cells and other viable but nontarget bacterial cells. Using recombinant M13KE-TC phage and Escherichia coli ER2738 as a model system, distinct differentiation between individual viable target cells from dead target cells was demonstrated by flow cytometry and fluorescence microscopy. As few as 1% viable E. coli ER2738 can be accurately quantified in a mix with dead E. coli ER2738 by flow cytometry. With fluorescence microscopic measurement, specific detection of as rare as 1 cfu/mL original viable target bacteria was achieved in the presence of a large excess of dead target cells and other viable but nontarget bacterial cells in 40 mL artificially contaminated drinking water sample in less than 3 h. This TC-phage-FlAsH approach is sensitive, specific, rapid, and simple, and thus shows great potential in water safety monitoring, health surveillance, and clinical diagnosis of which trace detection and identification of viable bacterial pathogens is highly demanded.

In vivo bacterial imaging without engineering; A novel probe-based strategy facilitated by endogenous nitroreductase enzymes.

PubMed

Stanton, Michael; Cronin, Michelle; Lehouritis, Panos; Tangney, Mark

2015-01-01

The feasibility of utilising bacteria as vectors for gene therapy is becoming increasingly recognised. This is primarily due to a number of intrinsic properties of bacteria such as their tumour targeting capabilities, their ability to carry large genetic or protein loads and the availability of well-established genetic engineering tools for a range of common lab strains. However, a number of issues relating to the use of bacteria as vectors for gene therapy need to be addressed in order for the field to progress. Amongst these is the need for the development of non-invasive detection/imaging systems for bacteria within a living host. In vivo optical imaging has advanced preclinical research greatly, and typically involves engineering of bacteria with genetic expression constructs for luminescence (e.g. the lux operon) or fluorescent proteins (GFP etc.). This requirement for genetic modification can be restrictive, where engineering is not experimentally appropriate or technologically feasible (e.g. due to lack of suitable engineering tools). We describe a novel strategy exploiting endogenous bacterial enzymatic activity to specifically activate an exogenously administered fluorescent imaging probe. The red shifted, quenched fluorophore CytoCy5S is reduced to a fluorescent form by bacterial-specific nitroreductase (NTR) enzymes. NTR enzymes are present in a wide range of bacterial genera and absent in mammalian systems, permitting highly specific detection of Gram-negative and Gram-positive bacteria in vivo. In this study, dose-responsive bacterial-specific signals were observed in vitro from all genera examined - E. coli, Salmonella, Listeria, Bifidobacterium and Clostridium difficile. Examination of an NTR-knockout strain validated the enzyme specificity of the probe. In vivo whole-body imaging permitted specific, dose-responsive monitoring of bacteria over time in various infection models, and no toxicity to bacteria or host was observed. This study demonstrates the concept of exploiting innate NTR activity as a reporting strategy for wild-type bacteria using optical imaging, while the concept may also be extended to NTR-specific probes for use with other imaging modalities.

Rationalizing and advancing the 3-MPBA SERS sandwich assay for rapid detection of bacteria in environmental and food matrices.

PubMed

Pearson, Brooke; Mills, Alexander; Tucker, Madeline; Gao, Siyue; McLandsborough, Lynne; He, Lili

2018-06-01

Bacterial foodborne illness continues to be a pressing issue in our food supply. Rapid detection methods are needed for perishable foods due to their short shelf lives and significant contribution to foodborne illness. Previously, a sensitive and reliable surface-enhanced Raman spectroscopy (SERS) sandwich assay based on 3-mercaptophenylboronic acid (3-MBPA) as a capturer and indicator molecule was developed for rapid bacteria detection. In this study, we explored the advantages and constraints of this assay over the conventional aerobic plate count (APC) method and further developed methods for detection in real environmental and food matrices. The SERS sandwich assay was able to detect environmental bacteria in pond water and on spinach leaves at higher levels than the APC method. In addition, the SERS assay appeared to have higher sensitivity to quantify bacteria in the stationary phase. On the other hand, the APC method was more sensitive to cell viability. Finally, a method to detect bacteria in a challenging high-sugar juice matrix was developed to enhance bacteria capture. This study advanced the SERS technique for real applications in environment and food matrices. Copyright © 2017 Elsevier Ltd. All rights reserved.

Absence of Bacteria on Coronary Angioplasty Balloons from Unselected Patients: Results with Use of a High Sensitivity Polymerase Chain Reaction Assay

PubMed Central

Hansen, Gorm Mørk; Nilsson, Martin; Nielsen, Claus Henrik; Holmstrup, Palle; Helqvist, Steffen; Tolker-Nielsen, Tim; Givskov, Michael; Hansen, Peter Riis

2015-01-01

Periodontitis is a chronic, bacterially-induced inflammatory disease of the tooth-supporting tissues, which may result in transient bacteremia and a systemic inflammatory response. Periodontitis is associated with coronary artery disease independently of established cardiovascular risk factors, and translocation of bacteria from the oral cavity to the coronary arteries may play a role in the development of coronary artery disease. Very few studies have used angioplasty balloons for in vivo sampling from diseased coronary arteries, and with varying results. Therefore, the aim of this study was to assess if bacterial DNA from primarily oral bacteria could be detected on coronary angioplasty balloons by use of an optimized sampling process combined with an internally validated sensitive polymerase chain reaction (PCR) assay. Coronary angioplasty balloons and control samples from a total of 45 unselected patients with stable angina, unstable angina/non-ST elevation myocardial infarction, and ST-elevation myocardial infarction (n = 15 in each group) were collected and analyzed using a PCR assay with high sensitivity and specificity for 16S rRNA genes of the oral microbiome. Despite elimination of extraction and purification steps, and demonstration of sensitivity levels of 25–125 colony forming units (CFU), we did not detect bacterial DNA from any of the coronary angioplasty balloons. A subsequent questionnaire indicated that the prevalence of periodontitis in the study cohort was at least 39.5%. Although coronary angioplasty balloons are unlikely to be useful for detection of bacteria with current PCR techniques in unselected patients with coronary artery disease, more studies are warranted to determine the extent to which bacteria contribute to atherosclerosis and its clinical manifestations and whether the presence of bacteria in the arteries is a transient phenomenon. PMID:26695491

Bacteria, viruses, and parasites in an intermittent stream protected from and exposed to pasturing cattle: prevalence, densities, and quantitative microbial risk assessment.

PubMed

Wilkes, G; Brassard, J; Edge, T A; Gannon, V; Jokinen, C C; Jones, T H; Neumann, N; Pintar, K D M; Ruecker, N; Schmidt, P J; Sunohara, M; Topp, E; Lapen, D R

2013-10-15

Over 3500 individual water samples, for 131 sampling times, targeting waterborne pathogens/fecal indicator bacteria were collected during a 7-year period from 4 sites along an intermittent stream running through a small livestock pasture system with and without cattle access-to-stream restriction measures. The study assessed the impact of cattle pasturing/riparian zone protection on: pathogen (bacterial, viral, parasite) occurrence, concentrations of fecal indicators, and quantitative microbial risk assessments (QMRA) of the risk of Cryptosporidium, Giardia and Escherichia coli O157:H7 infection in humans. Methodologies were developed to compute QMRA mean risks on the basis of water samples exhibiting potentially human infectious Cryptosporidium and E. coli based on genotyping Crytosporidium, and E. coli O157:H7 presence/absence information paired with enumerated E. coli. All Giardia spp. were considered infectious. No significant pasturing treatment effects were observed among pathogens, with the exception of Campylobacter spp. and E. coli O157:H7. Campylobacter spp. prevalence significantly decreased downstream through pasture treatments and E. coli O157:H7 was observed in a few instances in the middle of the unrestricted pasture. Densities of total coliform, fecal coliform, and E. coli reduced significantly downstream in the restricted pasture system, but not in the unrestricted system. Seasonal and flow conditions were associated with greater indicator bacteria densities, especially in the summer. Norovirus GII was detected at rates of 7-22% of samples for all monitoring sites, and rotavirus in 0-7% of samples for all monitoring sites; pasture treatment trends were not evident, however. Seasonal and stream flow variables (and their interactions) were relatively more important than pasture treatments for initially stratifying pathogen occurrence and higher fecal indicator bacteria densities. Significant positive associations among fecal indicator bacteria and Campylobacter spp. detection were observed. For QMRA, adjusting for the proportion of Cryptosporidium spp. detected that are infectious for humans reduces downstream risk estimates by roughly one order of magnitude. Using QMRA in this manner provides a more refined estimate of beneficial management practice effects on pathogen exposure risks to humans. Crown Copyright © 2013. Published by Elsevier Ltd. All rights reserved.

Template reporter bacteriophage platform and multiple bacterial detection assays based thereon

NASA Technical Reports Server (NTRS)

Goodridge, Lawrence (Inventor)

2007-01-01

The invention is a method for the development of assays for the simultaneous detection of multiple bacteria. A bacteria of interest is selected. A host bacteria containing plasmid DNA from a T even bacteriophage that infects the bacteria of interest is infected with T4 reporter bacteriophage. After infection, the progeny bacteriophage are plating onto the bacteria of interest. The invention also includes single-tube, fast and sensitive assays which utilize the novel method.

Highly selective colorimetric bacteria sensing based on protein-capped nanoparticles.

PubMed

Qiu, Suyan; Lin, Zhenyu; Zhou, Yaomin; Wang, Donggen; Yuan, Lijuan; Wei, Yihua; Dai, Tingcan; Luo, Linguang; Chen, Guonan

2015-02-21

A rapid and cost-effective colorimetric sensor has been developed for the detection of bacteria (Bacillus subtilis was selected as an example). The sensor was designed to rely on lysozyme-capped AuNPs with the advantages of effective amplification and high specificity. In the sensing system, lysozyme was able to bind strongly to Bacillus subtilis, which effectively induced a color change of the solution from light purple to purplish red. The lowest concentration of Bacillus subtilis detectable by the naked eye was 4.5 × 10(3) colony-forming units (CFU) mL(-1). Similar results were discernable from UV-Vis absorption measurements. A good specificity was observed through a statistical analysis method using the SPSS software (version 17.0). This simple colorimetric sensor may therefore be a rapid and specific method for a bacterial detection assay in complex samples.

A novel surface-enhanced Raman scattering (SERS) detection for natural gas exploration using methane-oxidizing bacteria.

PubMed

Liang, Weiwei; Chen, Qiao; Peng, Fang; Shen, Aiguo; Hu, Jiming

2018-07-01

Methane-oxidizing bacteria (MOB), a unique group of Gram-negative bacteria utilizing methane as a sole source of carbon and energy, have been proved to be a biological indicator for gas prospecting. Field and cultivation-free detection of MOB is important but still challenging in current microbial prospecting of oil and gas (MPOG) system. Herein, SERS was used for the first time to our knowledge to investigate two species of methanotrophs and four closely relevant bacteria that universally coexisted in the upper soil of natural gas. A special but very simple approach was utilized to make silver nanoparticles (Ag NPs) sufficiently contact with every single bacterial cell, and highly strong and distinct Raman signals free from any native fluorescence have been obtained, and successfully utilized for distinguishing MOB from other species. A more convincing multi-Raman criterion based on single Raman bands, and further the entire Raman spectrum in combination with statistical analysis (e.g., principal component analysis (PCA)), which were found capable of classifying MOB related bacterial cells in soil with an accuracy of 100%. This study therefore demonstrated sensitive and rapid SERS measurement technique accompanied by complete Raman database of various gas reservoirs related bacteria could aid field exploration of natural gas reservoir. Copyright © 2018 Elsevier B.V. All rights reserved.

Diversity of sulfate-reducing bacteria in a plant using deep geothermal energy

NASA Astrophysics Data System (ADS)

Alawi, Mashal; Lerm, Stephanie; Vetter, Alexandra; Wolfgramm, Markus; Seibt, Andrea; Würdemann, Hilke

2011-06-01

Enhanced process understanding of engineered geothermal systems is a prerequisite to optimize plant reliability and economy. We investigated microbial, geochemical and mineralogical aspects of a geothermal groundwater system located in the Molasse Basin by fluid analysis. Fluids are characterized by temperatures ranging from 61°C to 103°C, salinities from 600 to 900 mg/l and a dissolved organic carbon content (DOC) between 6.4 to 19.3 mg C/l. The microbial population of fluid samples was analyzed by genetic fingerprinting techniques based on PCR-amplified 16S rRNA- and dissimilatory sulfite reductase genes. Despite of the high temperatures, microbes were detected in all investigated fluids. Fingerprinting and DNA sequencing enabled a correlation to metabolic classes and biogeochemical processes. The analysis revealed a broad diversity of sulfate-reducing bacteria. Overall, the detection of microbes known to be involved in biocorrosion and mineral precipitation indicates that microorganisms could play an important role for the understanding of processes in engineered geothermal systems.

Occurrence of fecal-indicator bacteria and protocols for identification of fecal-contamination sources in selected reaches of the West Branch Brandywine Creek, Chester County, Pennsylvania

USGS Publications Warehouse

Cinotto, Peter J.

2005-01-01

The presence of fecal-indicator bacteria indicates the potential presence of pathogens originating from the fecal matter of warm-blooded animals. These pathogens are responsible for numerous human diseases ranging from common diarrhea to meningitis and polio. The detection of fecal-indicator bacteria and interpretation of the resultant data are, therefore, of great importance to water-resource managers. Current (2005) techniques used to assess fecal contamination within the fluvial environment primarily assess samples collected from the water column, either as grab samples or as depth- and (or) width-integrated samples. However, current research indicates approximately 99 percent of all bacteria within nature exist as attached, or sessile, bacteria. Because of this condition, most current techniques for the detection of fecal contamination, which utilize bacteria, assess only about 1 percent of the total bacteria within the fluvial system and are, therefore, problematic. Evaluation of the environmental factors affecting the occurrence and distribution of bacteria within the fluvial system, as well as the evaluation and modification of alternative approaches that effectively quantify the larger population of sessile bacteria within fluvial sediments, will present water-resource managers with more effective tools to assess, prevent, and (or) eliminate sources of fecal contamination within pristine and impaired watersheds. Two stream reaches on the West Branch Brandywine Creek in the Coatesville, Pa., region were studied between September 2002 and August 2003. The effects of sediment particle size, climatic conditions, aquatic growth, environmental chemistry, impervious surfaces, sediment and soil filtration, and dams on observed bacteria concentrations were evaluated. Alternative approaches were assessed to better detect geographic sources of fecal contamination including the use of turbidity as a surrogate for bacteria, the modification and implementation of sandbag bacteria samplers, and the use of optical brighteners. For the purposes of this report, sources of bacteria were defined as geographic locations where elevated concentrations of bacteria are observed within, or expected to enter, the main branch of the West Branch Brandywine Creek. Biologic sources (for example, waterfowl) were noted where applicable; however, no specific study of biologic sources (such as bacterial source tracking) was conducted. Data indicated that specific bacterial populations within fluvial sediments could be related to specific particle-size ranges. This relation is likely the result of the reduced porosity and permeability associated with finer sediments and the ability of specific bacteria to tolerate particular environments. Escherichia coli (E. coli) showed a higher median concentration (2,160 colonies per gram of saturated sediment) in the 0.125 to 0.5-millimeter size range of natural sediments than in other ranges, and enterococcus bacteria showed a higher median concentration (61,830 colonies per gram of saturated sediment) in the 0.062 to 0.25-millimeter size range of natural sediments than in other ranges. There were insufficient data to assess the particle-size relation to fecal coliform bacteria and (or) fecal streptococcus bacteria. Climatic conditions were shown to affect bacteria concentrations in both the water column and fluvial sediments. Drought conditions in 2002 resulted in lower overall bacteria concentrations than the more typically wet year of 2003. E. coli concentrations in fluvial sediment along the Coatesville study reach in 2002 had a median concentration of 92 colonies per gram of saturated sediment; in 2003, the median concentration had risen to 4,752 colonies per gram of saturated sediment. Symbiotic relations between bacteria and aquatic growth were likely responsible for increased bacteria concentrations observed within an impoundment area on the Coatesville study reach. This reach showed evidence of

[Bactericidal effect of soybean peroxidase-hydrogen peroxide-potassium iodide system].

PubMed

Jin, Jianling; Zhang, Weican; Li, Yu; Zhao, Yue; Wang, Fei; Gao, Peiji

2011-03-01

To study the bactericidal effect and the possible mechanisms of the three components system [soybean peroxidases (SBP)-hydrogen peroxide (H2O2)-potassium iodide (KI), SBP-H2O2-KI]. The inhibition and bactericidal effect of SBP-H2O2-KI system to bacteria was detected by OD600 and the number of live bacteria (CFU). The sensitivity was tested by comparing the minimum inhibitory concentration (MIC) of bacterial cultures before and after cultured under sub-lethal dose of SBP-H2O2-KI system. Oxidizing activity groups were detected with physical and chemical methods in order to explain the bactericidal mechanisms of SBP-H2O2-KI system. SBP-H2O2-KI ternary system had rapid and high efficient bactericidal effect to a variety of bacterial strains in just several minutes. The MICs had no significant changes when bacterial cultures continuously cultured in sub-lethal dose of SBP-H2O2-KI system, and no resistance/tolerance mutant strains could be isolated from them. Both physical and chemical test results showed that no hydroxyl radical produced in SBP- H2O2-KI reaction system, chemical test results showed that no superoxide anion but a singlet oxygen and iodine produced in SBP-H2O2-KI reaction system. These results suggested that singlet oxygen and iodine or the iodine intermediate state may possible be the main sterilization factors for SBP-H2O2-KI system, and hydroxyl radical and superoxide anion not. In addition, the both characteristics of SBP-H2O2-KI system: rapid and high efficient bactericidal effect, and bacteria difficultly resisting to it, indicated it would have a good potential application in medical and plant protection area.

Magnetic biosensor using a high transition temperature SQUID

NASA Astrophysics Data System (ADS)

Grossman, Helene Lila

A high transition temperature (Tc) Superconducting QUantum Interference Device (SQUID) is used to detect magnetically-labeled microorganisms. The targets are identified and quantified by means of magnetic relaxation measurements, with no need for unbound magnetic labels to be washed away. The binding rate between antibody-linked magnetic particles and targets can be measured with this technique. Installed in a "SQUID microscope," a YBa2Cu 3O7-delta SQUID is mounted on a sapphire rod thermally linked to a liquid nitrogen can; these components are enclosed in a fiberglass vacuum chamber. A thin window separates the vacuum chamber from the sample, which is at room temperature and atmospheric pressure. In one mode of the experiment, targets are immobilized on a substrate and immersed a suspension of ˜50 nm diameter superparamagnetic particles, coated with antibodies. A pulsed magnetic field aligns the magnetic dipole moments, and the SQUID measures the magnetic relaxation signal each time the field is turned off. Unbound particles relax within ˜50 mus by Brownian rotation, too fast for the SQUID system to measure. In contrast, particles bound to targets have their Brownian motion inhibited. These particles relax in ˜1 s by rotation of the internal dipole moment, and this Neel relaxation process is detected by the SQUID. This assay is demonstrated with a model system of liposomes carrying the FLAG epitope; the detection limit is (2.7 +/- 0.2) x 105 particles. The replacement of the SQUID with a gradiometer improves the detection limit to (7.0 +/- 0.7) x 103 particles. In an alternate mode of the experiment, freely suspended targets (larger than ˜1 mum diameter) are detected. Since the Brownian relaxation time of the targets is longer than the measurement time, particles bound to targets are effectively immobilized and exhibit Neel relaxation. Listeria monocytogenes are detected using this method; the sensitivity is (1.1 +/- 0.2) x 105 bacteria in 20 muL. For a 1 nL sample volume, the detection limit is expected to be 230 +/- 40 bacteria. Time-resolved measurements, which yield the binding rate between particles and bacteria, are reported. Also, potential improvements to the system and possible applications are discussed.

Genome-wide detection of conservative site-specific recombination in bacteria

PubMed Central

Mathias Garrett, Elizabeth; Camilli, Andrew

2018-01-01

The ability of clonal bacterial populations to generate genomic and phenotypic heterogeneity is thought to be of great importance for many commensal and pathogenic bacteria. One common mechanism contributing to diversity formation relies on the inversion of small genomic DNA segments in a process commonly referred to as conservative site-specific recombination. This phenomenon is known to occur in several bacterial lineages, however it remains notoriously difficult to identify due to the lack of conserved features. Here, we report an easy-to-implement method based on high-throughput paired-end sequencing for genome-wide detection of conservative site-specific recombination on a single-nucleotide level. We demonstrate the effectiveness of the method by successfully detecting several novel inversion sites in an epidemic isolate of the enteric pathogen Clostridium difficile. Using an experimental approach, we validate the inversion potential of all detected sites in C. difficile and quantify their prevalence during exponential and stationary growth in vitro. In addition, we demonstrate that the master recombinase RecV is responsible for the inversion of some but not all invertible sites. Using a fluorescent gene-reporter system, we show that at least one gene from a two-component system located next to an invertible site is expressed in an on-off mode reminiscent of phase variation. We further demonstrate the applicability of our method by mining 209 publicly available sequencing datasets and show that conservative site-specific recombination is common in the bacterial realm but appears to be absent in some lineages. Finally, we show that the gene content associated with the inversion sites is diverse and goes beyond traditionally described surface components. Overall, our method provides a robust platform for detection of conservative site-specific recombination in bacteria and opens a new avenue for global exploration of this important phenomenon. PMID:29621238

Development of a biosensor protein bullet as a fluorescent method for fast detection of Escherichia coli in drinking water.

PubMed

Gutiérrez-Del-Río, Ignacio; Marín, Laura; Fernández, Javier; Álvarez San Millán, María; Ferrero, Francisco Javier; Valledor, Marta; Campo, Juan Carlos; Cobián, Natalia; Méndez, Ignacio; Lombó, Felipe

2018-01-01

Drinking water can be exposed to different biological contaminants from the source, through the pipelines, until reaching the final consumer or industry. Some of these are pathogenic bacteria and viruses which may cause important gastrointestinal or systemic diseases. The microbiological quality of drinking water relies mainly in monitoring three indicator bacteria of faecal origin, Escherichia coli, Enterococcus faecalis and Clostridium perfringens, which serve as early sentinels of potential health hazards for the population. Here we describe the analysis of three chimeric fluorescent protein bullets as biosensor candidates for fast detection of E. coli in drinking water. Two of the chimeric proteins (based on GFP-hadrurin and GFP-pb5 chimera proteins) failed with respect to specificity and/or sensitivity, but the GFP-colS4 chimera protein was able to carry out specific detection of E. coli in drinking water samples in a procedure encompassing about 8 min for final result and this biosensor protein was able to detect in a linear way between 20 and 103 CFU of this bacterium. Below 20 CFU, the system cannot differentiate presence or absence of the target bacterium. The fluorescence in this biosensor system is provided by the GFP subunit of the chimeric protein, which, in the case of the better performing sensor bullet, GFP-colS4 chimera, is covalently bound to a flexible peptide bridge and to a bacteriocin binding specifically to E. coli cells. Once bound to the target bacteria, the excitation step with 395 nm LED light causes emission of fluorescence from the GFP domain, which is amplified in a photomultiplier tube, and finally this signal is converted into an output voltage which can be associated with a CFU value and these data distributed along mobile phone networks, for example. This method, and the portable fluorimeter which has been developed for it, may contribute to reduce the analysis time for detecting E. coli presence in drinking water.

Development of a biosensor protein bullet as a fluorescent method for fast detection of Escherichia coli in drinking water

PubMed Central

Gutiérrez-del-Río, Ignacio; Marín, Laura; Fernández, Javier; Álvarez San Millán, María; Ferrero, Francisco Javier; Valledor, Marta; Campo, Juan Carlos; Cobián, Natalia; Méndez, Ignacio

2018-01-01

Drinking water can be exposed to different biological contaminants from the source, through the pipelines, until reaching the final consumer or industry. Some of these are pathogenic bacteria and viruses which may cause important gastrointestinal or systemic diseases. The microbiological quality of drinking water relies mainly in monitoring three indicator bacteria of faecal origin, Escherichia coli, Enterococcus faecalis and Clostridium perfringens, which serve as early sentinels of potential health hazards for the population. Here we describe the analysis of three chimeric fluorescent protein bullets as biosensor candidates for fast detection of E. coli in drinking water. Two of the chimeric proteins (based on GFP-hadrurin and GFP-pb5 chimera proteins) failed with respect to specificity and/or sensitivity, but the GFP-colS4 chimera protein was able to carry out specific detection of E. coli in drinking water samples in a procedure encompassing about 8 min for final result and this biosensor protein was able to detect in a linear way between 20 and 103 CFU of this bacterium. Below 20 CFU, the system cannot differentiate presence or absence of the target bacterium. The fluorescence in this biosensor system is provided by the GFP subunit of the chimeric protein, which, in the case of the better performing sensor bullet, GFP-colS4 chimera, is covalently bound to a flexible peptide bridge and to a bacteriocin binding specifically to E. coli cells. Once bound to the target bacteria, the excitation step with 395 nm LED light causes emission of fluorescence from the GFP domain, which is amplified in a photomultiplier tube, and finally this signal is converted into an output voltage which can be associated with a CFU value and these data distributed along mobile phone networks, for example. This method, and the portable fluorimeter which has been developed for it, may contribute to reduce the analysis time for detecting E. coli presence in drinking water. PMID:29304041

Validating tyrosinase homologue melA as a photoacoustic reporter gene for imaging Escherichia coli

NASA Astrophysics Data System (ADS)

Paproski, Robert J.; Li, Yan; Barber, Quinn; Lewis, John D.; Campbell, Robert E.; Zemp, Roger

2015-10-01

To understand the pathogenic processes for infectious bacteria, appropriate research tools are required for replicating and characterizing infections. Fluorescence and bioluminescence imaging have primarily been used to image infections in animal models, but optical scattering in tissue significantly limits imaging depth and resolution. Photoacoustic imaging, which has improved depth-to-resolution ratio compared to conventional optical imaging, could be useful for visualizing melA-expressing bacteria since melA is a bacterial tyrosinase homologue which produces melanin. Escherichia coli-expressing melA was visibly dark in liquid culture. When melA-expressing bacteria in tubes were imaged with a VisualSonics Vevo LAZR system, the signal-to-noise ratio of a 9× dilution sample was 55, suggesting that ˜20 bacteria cells could be detected with our system. Multispectral (680, 700, 750, 800, 850, and 900 nm) analysis of the photoacoustic signal allowed unmixing of melA-expressing bacteria from blood. To compare photoacoustic reporter gene melA (using Vevo system) with luminescent and fluorescent reporter gene Nano-lantern (using Bruker Xtreme In-Vivo system), tubes of bacteria expressing melA or Nano-lantern were submerged 10 mm in 1% Intralipid, spaced between <1 and 20 mm apart from each other, and imaged with the appropriate imaging modality. Photoacoustic imaging could resolve the two tubes of melA-expressing bacteria even when the tubes were less than 1 mm from each other, while bioluminescence and fluorescence imaging could not resolve the two tubes of Nano-lantern-expressing bacteria even when the tubes were spaced 10 mm from each other. After injecting 100-μL of melA-expressing bacteria in the back flank of a chicken embryo, photoacoustic imaging allowed visualization of melA-expressing bacteria up to 10-mm deep into the embryo. Photoacoustic signal from melA could also be separated from deoxy- and oxy-hemoglobin signal observed within the embryo and chorioallantoic membrane. Our results suggest that melA is a useful photoacoustic reporter gene for visualizing bacteria, and further work incorporating photoacoustic reporters into infectious bacterial strains is warranted.

Rapid detection of food pathogens using RNA aptamers-immobilized slide.

PubMed

Maeng, Jin-Soo; Kim, Namsoo; Kim, Chong-Tai; Han, Seung Ryul; Lee, Young Ju; Lee, Seong-Wook; Lee, Myung-Hyun; Cho, Yong-Jin

2012-07-01

The purpose of this study was to develop a simple and rapid detection system for foodborne bacteria, which consisted of an optical microscope and its slide chip with artificial antibodies, or RNA aptamers. From an RNA pool, three each RNA aptamers were built by the method of SELEX (systematic evolution of ligands by exponential enrichment) for components of cell wall, LPS (lipopolysaccharide) from E. coli O157:H7, teichoic acid from Staphylococcus aureus and a cell membrane protein of OmpC from Salmonella typhimurium, respectively. These aptamers were hybridized with thiol-conjugated 16 dT-linker molecules in order to be immobilized on silver surface which was, in advance, fabricated on glass slide, using a spin-coating method. To confirm that each aptamers retained its specific binding activities to their antigenic live bacteria, microscopic view of bound cells immobilized on silver film were observed. Furthermore, we observed the fluorescence-emitting bacteria-aptamer complex immobilized on silver film after adding RNA aptamers hybridized with fluorophore, FAM-conjugated 16 dT-linker molecules. As a result, the RNA aptamers-immobilized slide system developed in this study was a useful new tool to rapidly monitor individual food pathogens.

Nitrotoga-like bacteria are previously unrecognized key nitrite oxidizers in full-scale wastewater treatment plants.

PubMed

Lücker, Sebastian; Schwarz, Jasmin; Gruber-Dorninger, Christiane; Spieck, Eva; Wagner, Michael; Daims, Holger

2015-03-01

Numerous past studies have shown members of the genus Nitrospira to be the predominant nitrite-oxidizing bacteria (NOB) in nitrifying wastewater treatment plants (WWTPs). Only recently, the novel NOB 'Candidatus Nitrotoga arctica' was identified in permafrost soil and a close relative was enriched from activated sludge. Still, little is known about diversity, distribution and functional importance of Nitrotoga in natural and engineered ecosystems. Here we developed Nitrotoga 16S rRNA-specific PCR primers and fluorescence in situ hybridization (FISH) probes, which were applied to screen activated sludge samples from 20 full-scale WWTPs. Nitrotoga-like bacteria were detected by PCR in 11 samples and reached abundances detectable by FISH in seven sludges. They coexisted with Nitrospira in most of these WWTPs, but constituted the only detectable NOB in two systems. Quantitative FISH revealed that Nitrotoga accounted for nearly 2% of the total bacterial community in one of these plants, a number comparable to Nitrospira abundances in other WWTPs. Spatial statistics revealed that Nitrotoga coaggregated with ammonia-oxidizing bacteria, strongly supporting a functional role in nitrite oxidation. This activity was confirmed by FISH in combination with microradiography, which revealed nitrite-dependent autotrophic carbon fixation by Nitrotoga in situ. Correlation of the presence or absence with WWTP operational parameters indicated low temperatures as a main factor supporting high Nitrotoga abundances, although in incubation experiments these NOB remained active over an unexpected range of temperatures, and also at different ambient nitrite concentrations. In conclusion, this study demonstrates that Nitrotoga can be functionally important nitrite oxidizers in WWTPs and can even represent the only known NOB in engineered systems.

Radiometric Method for the Detection of Coliform Organisms in Water

PubMed Central

Bachrach, Uriel; Bachrach, Zelilah

1974-01-01

A new radiometric method for the detection of coliform bacteria in water has been described. The method is based on the release of 14CO2 from [14C]lactose by bacteria suspended in growth medium and incubated at 37 C. The evolved 14CO2 is trapped by hyamine hydroxide and counted in a liquid scintillation spectrometer. The method permits the detection of 1 to 10 organisms within 6 h of incubation. Coliform bacteria suspended in water for several days recover from starvation and may be quantitated by the proposed method. Bacteria from water samples may also be concentrated by filtration through membrane filters and detected by the radiometric assay. PMID:4605007

Multiplexed nanoplasmonic biosensor for one-step simultaneous detection of Chlamydia trachomatis and Neisseria gonorrhoeae in urine.

PubMed

Soler, Maria; Belushkin, Alexander; Cavallini, Andrea; Kebbi-Beghdadi, Carole; Greub, Gilbert; Altug, Hatice

2017-08-15

Development of rapid and multiplexed diagnostic tools is a top priority to address the current epidemic problem of sexually transmitted diseases. Here we introduce a novel nanoplasmonic biosensor for simultaneous detection of the two most common bacterial infections: Chlamydia trachomatis and Neisseria gonorrhoeae. Our plasmonic microarray is composed of gold nanohole sensor arrays that exhibit the extraordinary optical transmission (EOT), providing highly sensitive analysis in a label-free configuration. The integration in a microfluidic system and the precise immobilization of specific antibodies on the individual sensor arrays allow for selective detection and quantification of the bacteria in real-time. We achieved outstanding sensitivities for direct immunoassay of urine samples, with a limit of detection of 300 colony forming units (CFU)/mL for C. trachomatis and 1500CFU/mL for N. gonorrhoeae. The multiplexing capability of our biosensor was demonstrated by analyzing different urine samples spiked with either C. trachomatis or N. gonorrhoeae, and also containing both bacteria. We could successfully detect, identify and quantify the levels of the two bacteria in a one-step assay, without the need for DNA extraction or amplification techniques. This work opens up new possibilities for the implementation of point-of-care biosensors that enable fast, simple and efficient diagnosis of sexually transmitted infections. Copyright © 2017 The Authors. Published by Elsevier B.V. All rights reserved.

Highly sensitive and quantitative detection of rare pathogens through agarose droplet microfluidic emulsion PCR at the single-cell level.

PubMed

Zhu, Zhi; Zhang, Wenhua; Leng, Xuefei; Zhang, Mingxia; Guan, Zhichao; Lu, Jiangquan; Yang, Chaoyong James

2012-10-21

Genetic alternations can serve as highly specific biomarkers to distinguish fatal bacteria or cancer cells from their normal counterparts. However, these mutations normally exist in very rare amount in the presence of a large excess of non-mutated analogs. Taking the notorious pathogen E. coli O157:H7 as the target analyte, we have developed an agarose droplet-based microfluidic ePCR method for highly sensitive, specific and quantitative detection of rare pathogens in the high background of normal bacteria. Massively parallel singleplex and multiplex PCR at the single-cell level in agarose droplets have been successfully established. Moreover, we challenged the system with rare pathogen detection and realized the sensitive and quantitative analysis of a single E. coli O157:H7 cell in the high background of 100,000 excess normal K12 cells. For the first time, we demonstrated rare pathogen detection through agarose droplet microfluidic ePCR. Such a multiplex single-cell agarose droplet amplification method enables ultra-high throughput and multi-parameter genetic analysis of large population of cells at the single-cell level to uncover the stochastic variations in biological systems.

[Detection of CRSPR-Cas system in Streptococcus thermophiles].

PubMed

Li, Wan; Liang, Hongzhang; Zhang, Danqing; Wang, Nana; Tang, Yaru; Li, Bailiang; Huo, Guicheng

2016-04-14

We aimed to detect the CRSPR-Cas system of six Streptococcus thermophilus. Bioinformatics method was used to predict CRSPR-Cas system of nine S. thermophilus that published in National Center for Biotechnology Information. Four primers were designed according to the flanking sequences of standard strains and the CRISPR-Cas system of six S. thermophilus have been detected by PCR method. S. thermophilus S4 had a Cas9 gene, others all had Cas9 gene, Cas10 gene and Cas9* gene. In addition, 79 and KLDS3.0207 still had Cas3 gene. Signature genes amplification of CRSPR-Cas system could predict the type of CRSPR-Cas system in unsequenced strains, these findings will help establish the foundation for the study of CRSPR-Cas system in lactic acid bacteria.

Efficiency of a closed-coupled solar pasteurization system in treating roof harvested rainwater.

PubMed

Dobrowsky, P H; Carstens, M; De Villiers, J; Cloete, T E; Khan, W

2015-12-01

Many studies have concluded that roof harvested rainwater is susceptible to chemical and microbial contamination. The aim of the study was thus to conduct a preliminary investigation into the efficiency of a closed-coupled solar pasteurization system in reducing the microbiological load in harvested rainwater and to determine the change in chemical components after pasteurization. The temperature of the pasteurized tank water samples collected ranged from 55 to 57°C, 64 to 66°C, 72 to 74°C, 78 to 81°C and 90 to 91°C. Cations analyzed were within drinking water guidelines, with the exception of iron [195.59 μg/L (55°C)-170.1 μg/L (91°C)], aluminum [130.98 μg/L (78°C)], lead [12.81 μg/L (55°C)-13.2 μg/L (91°C)] and nickel [46.43 μg/L (55°C)-32.82 μg/L (78°C)], which were detected at levels above the respective guidelines in the pasteurized tank water samples. Indicator bacteria including, heterotrophic bacteria, Escherichia coli and total coliforms were reduced to below the detection limit at pasteurization temperatures of 72°C and above. However, with the use of molecular techniques Yersinia spp., Legionella spp. and Pseudomonas spp. were detected in tank water samples pasteurized at temperatures greater than 72°C. The viability of the bacteria detected in this study at the higher temperature ranges should thus be assessed before pasteurized harvested rainwater is used as a potable water source. In addition, it is recommended that the storage tank of the pasteurization system be constructed from an alternative material, other than stainless steel, in order for a closed-coupled pasteurization system to be implemented and produce large quantities of potable water from roof harvested rainwater. Copyright © 2015 Elsevier B.V. All rights reserved.

Microbial fouling community analysis of the cooling water system of a nuclear test reactor with emphasis on sulphate reducing bacteria.

PubMed

Balamurugan, P; Joshi, M Hiren; Rao, T S

2011-10-01

Culture and molecular-based techniques were used to characterize bacterial diversity in the cooling water system of a fast breeder test reactor (FBTR). Techniques were selected for special emphasis on sulphate-reducing bacteria (SRB). Water samples from different locations of the FBTR cooling water system, in addition to biofilm scrapings from carbon steel coupons and a control SRB sample were characterized. Whole genome extraction of the water samples and SRB diversity by group specific primers were analysed using nested PCR and denaturing gradient gel electrophoresis (DGGE). The results of the bacterial assay in the cooling water showed that the total culturable bacteria (TCB) ranged from 10(3) to 10(5) cfu ml(-1); iron-reducing bacteria, 10(3) to 10(5) cfu ml(-1); iron oxidizing bacteria, 10(2) to 10(3) cfu ml(-1) and SRB, 2-29 cfu ml(-1). However, the counts of the various bacterial types in the biofilm sample were 2-3 orders of magnitude higher. SRB diversity by the nested PCR-DGGE approach showed the presence of groups 1, 5 and 6 in the FBTR cooling water system; however, groups 2, 3 and 4 were not detected. The study demonstrated that the PCR protocol influenced the results of the diversity analysis. The paper further discusses the microbiota of the cooling water system and its relevance in biofouling.

Raman Hyperspectral Imaging for Detection of Watermelon Seeds Infected with Acidovorax citrulli.

PubMed

Lee, Hoonsoo; Kim, Moon S; Qin, Jianwei; Park, Eunsoo; Song, Yu-Rim; Oh, Chang-Sik; Cho, Byoung-Kwan

2017-09-23

The bacterial infection of seeds is one of the most important quality factors affecting yield. Conventional detection methods for bacteria-infected seeds, such as biological, serological, and molecular tests, are not feasible since they require expensive equipment, and furthermore, the testing processes are also time-consuming. In this study, we use the Raman hyperspectral imaging technique to distinguish bacteria-infected seeds from healthy seeds as a rapid, accurate, and non-destructive detection tool. We utilize Raman hyperspectral imaging data in the spectral range of 400-1800 cm -1 to determine the optimal band-ratio for the discrimination of watermelon seeds infected by the bacteria Acidovorax citrulli using ANOVA. Two bands at 1076.8 cm -1 and 437 cm -1 are selected as the optimal Raman peaks for the detection of bacteria-infected seeds. The results demonstrate that the Raman hyperspectral imaging technique has a good potential for the detection of bacteria-infected watermelon seeds and that it could form a suitable alternative to conventional methods.

Raman Hyperspectral Imaging for Detection of Watermelon Seeds Infected with Acidovorax citrulli

PubMed Central

Lee, Hoonsoo; Kim, Moon S.; Qin, Jianwei; Park, Eunsoo; Song, Yu-Rim; Oh, Chang-Sik

2017-01-01

The bacterial infection of seeds is one of the most important quality factors affecting yield. Conventional detection methods for bacteria-infected seeds, such as biological, serological, and molecular tests, are not feasible since they require expensive equipment, and furthermore, the testing processes are also time-consuming. In this study, we use the Raman hyperspectral imaging technique to distinguish bacteria-infected seeds from healthy seeds as a rapid, accurate, and non-destructive detection tool. We utilize Raman hyperspectral imaging data in the spectral range of 400–1800 cm−1 to determine the optimal band-ratio for the discrimination of watermelon seeds infected by the bacteria Acidovorax citrulli using ANOVA. Two bands at 1076.8 cm−1 and 437 cm−1 are selected as the optimal Raman peaks for the detection of bacteria-infected seeds. The results demonstrate that the Raman hyperspectral imaging technique has a good potential for the detection of bacteria-infected watermelon seeds and that it could form a suitable alternative to conventional methods. PMID:28946608

[Detection of endotoxins of Gram-negative bacteria on the basis of electromagnetic radiation frequency spectrum].

PubMed

Likhoded, V G; Kuleshova, N V; Sergieva, N V; Konev, Iu V; Trubnikova, I A; Sudzhian, E V

2007-01-01

Method of Gram-negative bacteria endotoxins detection on the basis of their own spectrum of electromagnetic radiation frequency was developed. Frequency spectrum typical for chemotype Re glycolipid, which is a part of lypopolysaccharides in the majority of Gram-negative bacteria, was used. Two devices--"Mini- Expert-DT" (manufactured by IMEDIS, Moscow) and "Bicom" (manufactured by Regumed, Germany)--were used as generators of electromagnetic radiation. Detection of endotoxin using these devices was performed by electropuncture vegetative resonance test. Immunoenzyme reaction with antibodies to chemotype Re glycolipid was used during analysis of preparations for assessment of resonance-frequency method specificity. The study showed that resonance-frequency method can detect lypopolysaccharides of different enterobacteria in quantities up to 0.1 pg as well as bacteria which contain lypopolysaccharides. At the same time, this method does not detect such bacteria as Staphylococcus aureus, Bifidobacterium spp., Lactobacillus spp., and Candida albicans. The method does not require preliminary processing of blood samples and can be used for diagnostics of endotoxinemia, and detection of endotoxins in blood samples or injection solutions.

Real-time PCR for rapidly detecting aniline-degrading bacteria in activated sludge.

PubMed

Kayashima, Takakazu; Suzuki, Hisako; Maeda, Toshinari; Ogawa, Hiroaki I

2013-05-01

We developed a detection method that uses quantitative real-time PCR (qPCR) and the TaqMan system to easily and rapidly assess the population of aniline-degrading bacteria in activated sludge prior to conducting a biodegradability test on a chemical compound. A primer and probe set for qPCR was designed by a multiple alignment of conserved amino acid sequences encoding the large (α) subunit of aniline dioxygenase. PCR amplification tests showed that the designed primer and probe set targeted aniline-degrading strains such as Acidovorax sp., Gordonia sp., Rhodococcus sp., and Pseudomonas putida, thereby suggesting that the developed method can detect a wide variety of aniline-degrading bacteria. There was a strong correlation between the relative copy number of the α-aniline dioxygenase gene in activated sludge obtained with the developed qPCR method and the number of aniline-degrading bacteria measured by the Most Probable Number method, which is the conventional method, and a good correlation with the lag time of the BOD curve for aniline degradation produced by the biodegradability test in activated sludge samples collected from eight different wastewater treatment plants in Japan. The developed method will be valuable for the rapid and accurate evaluation of the activity of inocula prior to conducting a ready biodegradability test. Copyright © 2013 Elsevier Ltd. All rights reserved.

[Trend survey of ocular infections with bacteria at Toyama University Hospital over the past six years--from the standpoint of laboratory examination].

PubMed

Kubota, Tomomi; Hayashi, Shirou; Niimi, Hideki; Kitajima, Isao

2012-07-01

Specimens of bacterial ocular infections are frequently received in the clinical laboratory. However, a comprehensive trend survey of ocular infections with bacteria is very rare. Our objective is to understand the current tendency of ocular infections with bacteria in patients at Toyama University Hospital from the standpoint of laboratory examination. We studied 263 cases of ocular infection with bacteria diagnosed at Toyama University Hospital from January 2006 to December 2011. 123 were male and 140 were female, with a mean age of 61.2(0-98) years. Specimens were subjected to direct microscopy and culture. Cultures were positive in 174(66.2%) patients. The most common bacterial isolate was Staphylococcus (28.1%), followed by Corynebacterium (19.3%), Streptococcus (9.3%), and Propionibacterium (8.6%). MRSA accounted for 18.8% of all S. aureus isolates, and has increased in recent years. The number of bacteria detected was larger in March, June, July, August, and October. Age distribution indicated that around 70% of bacterial isolates were detected from patients over 60 years old. The most common specimen of ocular infections with bacteria was eye discharge (detection rate; 87.8%), followed by corneal scraping(41%), aqueous humor (19%), and vitreous body (27%). Nearly 80% of bacterial isolates were detected from patients with keratitis, endophthalmitis, dacryocystitis, and conjunctivitis. As for the disease specific detection rate, endophthalmitis was very low (38.3%). The detection rate by years indicated that the way doctors pick up the specimens greatly affects the detection rate. Based on this survey, we need close cooperation with medical doctors concerning laboratory examination in ocular infection with bacteria, and we must improve the detection sensitivity of specimens from patients with endophthalmitis.

Microthrix parvicella and Gordona amarae in mesophilic and thermophilic anaerobic digestion systems.

PubMed

Marneri, Matina; Mamais, Daniel; Koutsiouki, Efi

2009-04-14

The scope of the study presented in this paper is to determine the fate of the filamentous bacteria Gordona amarae and Microthrix parvicella in anaerobic digestion operating under mesophilic and thermophilic conditions. In order to detect and quantify foaming bacteria in the anaerobic digesters, a fluorescent in situ hybridization (FISH) method was developed and applied. This paper presents the results of a laboratory-scale study that involved the operation of four lab-scale anaerobic digestion systems operating in the mesophilic (35 degrees C) and thermophilic (55 degrees C) temperature ranges at 20 days' detention time. According to the FISH counts of G. amarae and M. parvicella, it appears that thermophilic conditions resulted in a higher destruction of both filamentous bacteria, averaging approximately 97% and 94% for the single thermophilic digester and the dual thermophilic/mesophilic system, respectively. Within the context of this study, the overall performance of the four different anaerobic digestion systems was evaluated in terms of biogas production per mass of volatile solids destroyed, COD destruction, sludge dewaterability and foaming characteristics. The dual stage systems used in this study outperformed the single stage digesters.

Rapid, highly sensitive detection of Gram-negative bacteria with lipopolysaccharide based disposable aptasensor.

PubMed

Zhang, Jian; Oueslati, Rania; Cheng, Cheng; Zhao, Ling; Chen, Jiangang; Almeida, Raul; Wu, Jayne

2018-07-30

Gram-negative bacteria are one of the most common microorganisms in the environment. Their differential detection and recognition from Gram-positive bacteria has been attracting much attention over the years. Using Escherichia coli (E. coli) as a model, we demonstrated on-site detection of Gram-negative bacteria by an AC electrokinetics-based capacitive sensing method using commercial microelectrodes functionalized with an aptamer specific to lipopolysaccharides. Dielectrophoresis effect was utilized to enrich viable bacteria to the microelectrodes rapidly, achieving a detection limit of 10 2 cells/mL within a 30 s' response time. The sensor showed a negligible response to Staphylococcus aureus (S. aureus), a Gram-positive species. The developed sensor showed significant advantages in sensitivity, selectivity, cost, operation simplicity, and response time. Therefore, this sensing method has shown great application potential for environmental monitoring, food safety, and real-time diagnosis. Copyright © 2018 Elsevier B.V. All rights reserved.

Differentiating the growth phases of single bacteria using Raman spectroscopy

NASA Astrophysics Data System (ADS)

Strola, S. A.; Marcoux, P. R.; Schultz, E.; Perenon, R.; Simon, A.-C.; Espagnon, I.; Allier, C. P.; Dinten, J.-M.

2014-03-01

In this paper we present a longitudinal study of bacteria metabolism performed with a novel Raman spectrometer system. Longitudinal study is possible with our Raman setup since the overall procedure to localize a single bacterium and collect a Raman spectrum lasts only 1 minute. Localization and detection of single bacteria are performed by means of lensfree imaging, whereas Raman signal (from 600 to 3200 cm-1) is collected into a prototype spectrometer that allows high light throughput (HTVS technology, Tornado Spectral System). Accomplishing time-lapse Raman spectrometry during growth of bacteria, we observed variation in the net intensities for some band groups, e.g. amides and proteins. The obtained results on two different bacteria species, i.e. Escherichia coli and Bacillus subtilis clearly indicate that growth affects the Raman chemical signature. We performed a first analysis to check spectral differences and similarities. It allows distinguishing between lag, exponential and stationary growth phases. And the assignment of interest bands to vibration modes of covalent bonds enables the monitoring of metabolic changes in bacteria caused by growth and aging. Following the spectra analysis, a SVM (support vector machine) classification of the different growth phases is presented. In sum this longitudinal study by means of a compact and low-cost Raman setup is a proof of principle for routine analysis of bacteria, in a real-time and non-destructive way. Real-time Raman studies on metabolism and viability of bacteria pave the way for future antibiotic susceptibility testing.

Gut dysbiosis and detection of "live gut bacteria" in blood of Japanese patients with type 2 diabetes.

PubMed

Sato, Junko; Kanazawa, Akio; Ikeda, Fuki; Yoshihara, Tomoaki; Goto, Hiromasa; Abe, Hiroko; Komiya, Koji; Kawaguchi, Minako; Shimizu, Tomoaki; Ogihara, Takeshi; Tamura, Yoshifumi; Sakurai, Yuko; Yamamoto, Risako; Mita, Tomoya; Fujitani, Yoshio; Fukuda, Hiroshi; Nomoto, Koji; Takahashi, Takuya; Asahara, Takashi; Hirose, Takahisa; Nagata, Satoru; Yamashiro, Yuichiro; Watada, Hirotaka

2014-08-01

Mounting evidence indicates that the gut microbiota are an important modifier of obesity and diabetes. However, so far there is no information on gut microbiota and "live gut bacteria" in the systemic circulation of Japanese patients with type 2 diabetes. Using a sensitive reverse transcription-quantitative PCR (RT-qPCR) method, we determined the composition of fecal gut microbiota in 50 Japanese patients with type 2 diabetes and 50 control subjects, and its association with various clinical parameters, including inflammatory markers. We also analyzed the presence of gut bacteria in blood samples. The counts of the Clostridium coccoides group, Atopobium cluster, and Prevotella (obligate anaerobes) were significantly lower (P < 0.05), while the counts of total Lactobacillus (facultative anaerobes) were significantly higher (P < 0.05) in fecal samples of diabetic patients than in those of control subjects. Especially, the counts of Lactobacillus reuteri and Lactobacillus plantarum subgroups were significantly higher (P < 0.05). Gut bacteria were detected in blood at a significantly higher rate in diabetic patients than in control subjects (28% vs. 4%, P < 0.01), and most of these bacteria were Gram-positive. This is the first report of gut dysbiosis in Japanese patients with type 2 diabetes as assessed by RT-qPCR. The high rate of gut bacteria in the circulation suggests translocation of bacteria from the gut to the bloodstream. © 2014 by the American Diabetes Association. Readers may use this article as long as the work is properly cited, the use is educational and not for profit, and the work is not altered.

Coliform inhibition by bacteriocin-like substances in drinking water distribution systems.

PubMed Central

Means, E G; Olson, B H

1981-01-01

Bacterial isolates from an unchlorinated potable groundwater system and a chlorinated surface water system were screened by an agar overlay method for the ability to produce bacteriocin-like substances (BLS) inhibitory to the growth of Escherichia coli, Klebsiella sp., and Enterobacter aerogenes. The production of coliform-specific BLS by noncoliform bacteria varied with the site and date of isolation as well as the genus of the producer strain. A total of 448 bacterial isolates were screened from the chlorinated system, and 22.1% produced BLS specific for at least one of the three coliforms. In the unchlorinated system, 7.9% (n = 696) possessed this ability. Flavobacterium/Moraxella comprised 57.1% of all bacteria (from both systems) producing BLS. The possibility that BLS interfere with coliform detection in standard bacteriological water quality tests is discussed. Images PMID:7027953

Comparison of Clark's presence-absence test and the membrane filter method for coliform detection in potable water samples.

PubMed Central

Pipes, W O; Minnigh, H A; Moyer, B; Troy, M A

1986-01-01

A total of 2,601 water samples from six different water systems were tested for coliform bacteria by Clark's presence-absence (P-A) test and by the membrane filter (MF) method. There was no significant difference in the fraction of samples positive for coliform bacteria for any of the systems tested. It was concluded that the two tests are equivalent for monitoring purposes. However, 152 samples were positive for coliform bacteria by the MF method but negative by the P-A test, and 132 samples were positive by the P-A test but negative by the MF method. Many of these differences for individual samples can be explained by random dispersion of bacteria in subsamples when the coliform density is low. However, 15 samples had MF counts greater than 3 and gave negative P-A results. The only apparent explanation for most of these results is that coliform bacteria were present in the P-A test bottles but did not produce acid and gas. Two other studies have reported more samples positive by Clark's P-A test than by the MF method. PMID:3532953

Influence of Chicken Manure Fertilization on Antibiotic-Resistant Bacteria in Soil and the Endophytic Bacteria of Pakchoi.

PubMed

Yang, Qingxiang; Zhang, Hao; Guo, Yuhui; Tian, Tiantian

2016-06-30

Animal manure is commonly used as fertilizer for agricultural crops worldwide, even though it is believed to contribute to the spread of antibiotic resistance from animal intestines to the soil environment. However, it is unclear whether and how there is any impact of manure fertilization on populations and community structure of antibiotic-resistant endophytic bacteria (AREB) in plant tissues. To investigate the effect of manure and organic fertilizer on endophytic bacterial communities, pot experiments were performed with pakchoi grown with the following treatments: (1) non-treated; (2) chicken manure-treated and (3) organic fertilizer-treated. Manure or organic fertilizer significantly increased the abundances of total cultivable endophytic bacteria (TCEB) and AREB in pakchoi, and the effect of chicken manure was greater than that of organic fertilizer. Further, 16S rDNA sequencing and the phylogenetic analysis indicated that chicken manure or organic fertilizer application increased the populations of multiple antibiotic-resistant bacteria (MARB) in soil and multiple antibiotic-resistant endophytic bacteria (MAREB) in pakchoi. The identical multiple antibiotic-resistant bacterial populations detected in chicken manure, manure- or organic fertilizer-amended soil and the vegetable endophytic system were Brevundimonas diminuta, Brachybacterium sp. and Bordetella sp., suggesting that MARB from manure could enter and colonize the vegetable tissues through manure fertilization. The fact that some human pathogens with multiple antibiotic resistance were detected in harvested vegetables after growing in manure-amended soil demonstrated a potential threat to human health.

Influence of Chicken Manure Fertilization on Antibiotic-Resistant Bacteria in Soil and the Endophytic Bacteria of Pakchoi

PubMed Central

Yang, Qingxiang; Zhang, Hao; Guo, Yuhui; Tian, Tiantian

2016-01-01

Animal manure is commonly used as fertilizer for agricultural crops worldwide, even though it is believed to contribute to the spread of antibiotic resistance from animal intestines to the soil environment. However, it is unclear whether and how there is any impact of manure fertilization on populations and community structure of antibiotic-resistant endophytic bacteria (AREB) in plant tissues. To investigate the effect of manure and organic fertilizer on endophytic bacterial communities, pot experiments were performed with pakchoi grown with the following treatments: (1) non-treated; (2) chicken manure-treated and (3) organic fertilizer-treated. Manure or organic fertilizer significantly increased the abundances of total cultivable endophytic bacteria (TCEB) and AREB in pakchoi, and the effect of chicken manure was greater than that of organic fertilizer. Further, 16S rDNA sequencing and the phylogenetic analysis indicated that chicken manure or organic fertilizer application increased the populations of multiple antibiotic-resistant bacteria (MARB) in soil and multiple antibiotic-resistant endophytic bacteria (MAREB) in pakchoi. The identical multiple antibiotic-resistant bacterial populations detected in chicken manure, manure- or organic fertilizer-amended soil and the vegetable endophytic system were Brevundimonas diminuta, Brachybacterium sp. and Bordetella sp., suggesting that MARB from manure could enter and colonize the vegetable tissues through manure fertilization. The fact that some human pathogens with multiple antibiotic resistance were detected in harvested vegetables after growing in manure-amended soil demonstrated a potential threat to human health. PMID:27376311

Microbes in mercury-enriched geothermal springs in western North America.

PubMed

Geesey, Gill G; Barkay, Tamar; King, Sue

2016-11-01

Because geothermal environments contain mercury (Hg) from natural sources, microorganisms that evolved in these systems have likely adapted to this element. Knowledge of the interactions between microorganisms and Hg in geothermal systems may assist in understanding the long-term evolution of microbial adaptation to Hg with relevance to other environments where Hg is introduced from anthropogenic sources. A number of microbiological studies with supporting geochemistry have been conducted in geothermal systems across western North America. Approximately 1 in 5 study sites include measurements of Hg. Of all prokaryotic taxa reported across sites with microbiological and accompanying physicochemical data, 42% have been detected at sites in which Hg was measured. Genes specifying Hg reduction and detoxification by microorganisms were detected in a number of hot springs across the region. Archaeal-like sequences, representing two crenarchaeal orders and one order each of the Euryarchaeota and Thaumarchaeota, dominated in metagenomes' MerA (the mercuric reductase protein) inventories, while bacterial homologs were mostly found in one deeply sequenced metagenome. MerA homologs were more frequently found in metagenomes of microbial communities in acidic springs than in circumneutral or high pH geothermal systems, possibly reflecting higher bioavailability of Hg under acidic conditions. MerA homologs were found in hot springs prokaryotic isolates affiliated with Bacteria and Archaea taxa. Acidic sites with high Hg concentrations contain more of Archaea than Bacteria taxa, while the reverse appears to be the case in circumneutral and high pH sites with high Hg concentrations. However, MerA was detected in only a small fraction of the Archaea and Bacteria taxa inhabiting sites containing Hg. Nevertheless, the presence of MerA homologs and their distribution patterns in systems, in which Hg has yet to be measured, demonstrates the potential for detoxification by Hg reduction in these geothermal systems, particularly the low pH springs that are dominated by Archaea. Copyright © 2016 Elsevier B.V. All rights reserved.

Rapid detection of bacteria with miniaturized pyrolysis-gas chromatographic analysis

NASA Astrophysics Data System (ADS)

Mowry, Curtis; Morgan, Catherine H.; Baca, Quentin; Manginell, Ronald P.; Kottenstette, Richard J.; Lewis, Patrick; Frye-Mason, Gregory C.

2002-02-01

Rapid detection and identification of bacteria and other pathogens is important for many civilian and military applications. The profiles of biological markers such as fatty acids can be used to characterize biological samples or to distinguish bacteria at the gram-type, genera, and even species level. Common methods for whole cell bacterial analysis are neither portable nor rapid, requiring lengthy, labor intensive sample preparation and bench-scale instrumentation. These methods chemically derivatize fatty acids to produce more volatile fatty acid methyl esters (FAMEs) that can be separated and analyzed by a gas chromatograph (GC)/mass spectrometer. More recent publications demonstrate decreased sample preparation time with in situ derivatization of whole bacterial samples using pyrolysis/derivatization. Ongoing development of miniaturized pyrolysis/GC instrumentation by this department capitalizes on Sandia advances in the field of microfabricated chemical analysis systems ((mu) ChemLab). Microdevices include rapidly heated stages capable of pyrolysis or sample concentration, gas chromatography columns, and surface acoustic wave (SAW) sensor arrays. We will present results demonstrating the capabilities of these devices toward fulfilling the goal of portable, rapid detection and early warning of the presence of pathogens in air or water.

Detection of five potentially periodontal pathogenic bacteria in peri-implant disease: A comparison of PCR and real-time PCR.

PubMed

Schmalz, Gerhard; Tsigaras, Sandra; Rinke, Sven; Kottmann, Tanja; Haak, Rainer; Ziebolz, Dirk

2016-07-01

The aim of this study was to compare the microbial analysis methods of polymerase chain reaction (PCR) and real-time PCR (RT-PCR) in terms of detection of five selected potentially periodontal pathogenic bacteria in peri-implant disease. Therefore 45 samples of healthy, mucositis and peri-implantitis (n = 15 each) were assessed according to presence of the following bacteria using PCR (DNA-strip technology) and RT-PCR (fluorescent dye SYBR green-system): Aggregatibacter actinomycetemcomitans (Aa), Porphyromonas gingivalis (Pg), Treponema denticola (Td), Tanerella forsythia (Tf), and Fusobacterium nucleatum (Fn). There were no significant correlations between the bacterial and disease patterns, so the benefit of using microbiological tests for the diagnosis of peri-implant diseases is questionable. Correlations between the methods were highest for Tf (Kendall's Tau: 0.65, Spearman: 0.78), Fn (0.49, 0.61) and Td (0.49, 0.59). For Aa (0.38, 0.42) and Pg (0.04, 0.04), lower correlation values were detected. Accordingly, conventional semi-quantitative PCR seems to be sufficient for analyzing potentially periodontal pathogenic bacterial species. Copyright © 2016 Elsevier Inc. All rights reserved.

Detection of pathogenic gram negative bacteria using infrared thermography

NASA Astrophysics Data System (ADS)

Lahiri, B. B.; Divya, M. P.; Bagavathiappan, S.; Thomas, Sabu; Philip, John

2012-11-01

Detection of viable bacteria is of prime importance in all fields of microbiology and biotechnology. Conventional methods of enumerating bacteria are often time consuming and labor-intensive. All living organisms generate heat due to metabolic activities and hence, measurement of heat energy is a viable tool for detection and quantification of bacteria. In this article, we employ a non-contact and real time method - infrared thermography (IRT) for measurement of temperature variations in four clinically significant gram negative pathogenic bacteria, viz. Vibrio cholerae, Vibrio mimicus, Proteus mirabilis and Pseudomonas aeruginosa. We observe that, the energy content, defined as the ratio of heat generated by bacterial metabolic activities to the heat lost from the liquid medium to the surrounding, vary linearly with the bacterial concentration in all the four pathogenic bacteria. The amount of energy content observed in different species is attributed to their metabolisms and morphologies that affect the convection velocity and hence heat transport in the medium.

A Palaeozoic Puzzle in Cenozoic Science.

ERIC Educational Resources Information Center

Mikkelsen, Tom

1982-01-01

The sword-tailed horseshoe crab (Limulus polyphemus) has developed its own defense against bacteria surrounding it. This defense system, under the name "Limulus test," now provides medicine and hygiene with a valuable means of detecting bacterial endotoxins at extremely low levels. (Author/JN)

[Application of anaerobic bacteria detection in oral and maxillofacial infection].

PubMed

Bao, Zhen-ying; Lin, Qin; Meng, Yan-hong; He, Chun; Su, Jia-zeng; Peng, Xin

2016-02-18

To investigate the distribution and drug resistance of anaerobic bacteria in the patients with oral and maxillofacial infection. Aerobic and anaerobic bacteria cultures from 61 specimens of pus from the patients with oral and maxillofacial infection in the Department of Oral and Maxillofacial Surgery, Peking University School of Stomatology were identified. The culture type was evaluated by API 20A kit and drug resistance test was performed by Etest method. The clinical data and antibacterial agents for the treatment of the 61 cases were collected, and the final outcomes were recorded. The bacteria cultures were isolated from all the specimens, with aerobic bacteria only in 6 cases (9.8%), anaerobic bacteria only in 7 cases (11.5%), and both aerobic and anaerobic bacteria in 48 cases (78.7%). There were 55 infected cases (90.2%) with anaerobic bacteria, and 81 anaerobic bacteria stains were isolated. The highest bacteria isolation rate of Gram positive anaerobic bacteria could be found in Peptostreptococcus, Bifidobacterium and Pemphigus propionibacterium. No cefoxitin, amoxicillin/carat acid resistant strain was detected in the above three Gram positive anaerobic bacteria. The highest bacteria isolation rate of Gram negative anaerobic bacteria could be detected in Porphyromonas and Prevotella. No metronidazole, cefoxitin, amoxicillin/carat acid resistant strain was found in the two Gram negative anaerobic bacteria. In the study, 48 patients with oral and maxillofacial infection were treated according to the results of drug resistance testing, and the clinical cure rate was 81.3%. Mixed aerobic and anaerobic bacteria cultures are very common in most oral and maxillofacial infection patients. Anaerobic bacteria culture and drug resistance testing play an important role in clinical treatment.

75 FR 32295 - Expedited Approval of Alternative Test Procedures for the Analysis of Contaminants Under the Safe...

Federal Register 2010, 2011, 2012, 2013, 2014

2010-06-08

... Identification of Coliform Bacteria and Escherichia coli in Finished Waters, January 2007, Version 1.1... Membrane Filter Test Method for Detection and Identification of Coliform Bacteria and Escherichia coli in... Detection and Identification of Coliform Bacteria and Escherichia coli in Finished Waters. November, 2000...

Detection of antimicrobial resistance-associated proteins by titanium dioxide-facilitated intact bacteria mass spectrometry† †Electronic supplementary information (ESI) available. See DOI: 10.1039/c7sc04089j

PubMed Central

Zhu, Yingdi; Gasilova, Natalia; Jović, Milica; Qiao, Liang; Liu, Baohong; Lovey, Lysiane Tissières; Pick, Horst

2018-01-01

Titanium dioxide-modified target plates were developed to enhance intact bacteria analysis by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. The plates were designed to photocatalytically destroy the bacterial envelope structure and improve the ionization efficiency of intracellular components, thereby promoting the measurable mass range and the achievable detection sensitivity. Accordingly, a method for rapid detection of antimicrobial resistance-associated proteins, conferring bacterial resistance against antimicrobial drugs, was established by mass spectrometric fingerprinting of intact bacteria without the need for any sample pre-treatment. With this method, the variations in resistance proteins’ expression levels within bacteria were quickly measured from the relative peak intensities. This approach of resistance protein detection directly from intact bacteria by mass spectrometry is useful for fast discrimination of antimicrobial-resistant bacteria from their non-resistant counterparts whilst performing species identification. Also, it could be used as a rapid and convenient way for initial determination of the underlying resistance mechanisms. PMID:29719694

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ogura, Toshihiko, E-mail: t-ogura@aist.go.jp

Highlights: • We developed a high-sensitive frequency transmission electric-field (FTE) system. • The output signal was highly enhanced by applying voltage to a metal layer on SiN. • The spatial resolution of new FTE method is 41 nm. • New FTE system enables observation of the intact bacteria and virus in water. - Abstract: The high-resolution structural analysis of biological specimens by scanning electron microscopy (SEM) presents several advantages. Until now, wet bacterial specimens have been examined using atmospheric sample holders. However, images of unstained specimens in water using these holders exhibit very poor contrast and heavy radiation damage. Recently,more » we developed the frequency transmission electric-field (FTE) method, which facilitates the SEM observation of biological specimens in water without radiation damage. However, the signal detection system presents low sensitivity. Therefore, a high EB current is required to generate clear images, and thus reducing spatial resolution and inducing thermal damage to the samples. Here a high-sensitivity detection system is developed for the FTE method, which enhances the output signal amplitude by hundredfold. The detection signal was highly enhanced when voltage was applied to the metal layer on silicon nitride thin film. This enhancement reduced the EB current and improved the spatial resolution as well as the signal-to-noise ratio. The spatial resolution of a high-sensitive FTE system is 41 nm, which is considerably higher than previous FTE system. New FTE system can easily be utilised to examine various unstained biological specimens in water, such as living bacteria and viruses.« less

The Performance of the Four Anaerobic Blood Culture Bottles BacT/ALERT-FN, -FN Plus, BACTEC-Plus and -Lytic in Detection of Anaerobic Bacteria and Identification by Direct MALDI-TOF MS

PubMed Central

Almuhayawi, Mohammed; Altun, Osman; Abdulmajeed, Adam Dilshad; Ullberg, Måns; Özenci, Volkan

2015-01-01

Detection and identification of anaerobic bacteria in blood cultures (BC) is a well-recognized challenge in clinical microbiology. We studied 100 clinical anaerobic BC isolates to evaluate the performance of BacT/ALERT-FN, -FN Plus (BioMérieux), BACTEC-Plus and -Lytic (Becton Dickinson BioSciences) BC bottles in detection and time to detection (TTD) of anaerobic bacteria. BACTEC Lytic had higher detection rate (94/100, 94%) than BacT/ALERT FN Plus (80/100, 80%) (p<0.01) in the studied material. There was no significant difference in detection of anaerobic bacteria among the remaining bottle types. The 67 anaerobic bacteria that signalled positive in all four bottle types were analyzed to compare the time to detection (TTD) and isolates were directly identified by MALDI-TOF MS. There was a significant difference in TTD among the four bottle types (p<0.0001). The shortest median TTD was 18 h in BACTEC Lytic followed by BacT/ALERT FN (23.5 h), BACTEC Plus (27 h) and finally BacT/ALERT FN Plus (38 h) bottles. In contrast, MALDI-TOF MS performed similarly in all bottle types with accurate identification in 51/67 (76%) BacT/ALERT FN, 51/67 (76%) BacT/ALERT FN Plus, 53/67 (79%) BACTEC Plus and 50/67 (75%) BACTEC Lytic bottles. In conclusion, BACTEC Lytic bottles have significantly better detection rates and shorter TTD compared to the three other bottle types. The anaerobic BC bottles are equally suitable for direct MALDI-TOF MS for rapid and reliable identification of common anaerobic bacteria. Further clinical studies are warranted to investigate the performance of anaerobic BC bottles in detection of anaerobic bacteria and identification by direct MALDI-TOF MS. PMID:26554930

The Performance of the Four Anaerobic Blood Culture Bottles BacT/ALERT-FN, -FN Plus, BACTEC-Plus and -Lytic in Detection of Anaerobic Bacteria and Identification by Direct MALDI-TOF MS.

PubMed

Almuhayawi, Mohammed; Altun, Osman; Abdulmajeed, Adam Dilshad; Ullberg, Måns; Özenci, Volkan

2015-01-01

Detection and identification of anaerobic bacteria in blood cultures (BC) is a well-recognized challenge in clinical microbiology. We studied 100 clinical anaerobic BC isolates to evaluate the performance of BacT/ALERT-FN, -FN Plus (BioMérieux), BACTEC-Plus and -Lytic (Becton Dickinson BioSciences) BC bottles in detection and time to detection (TTD) of anaerobic bacteria. BACTEC Lytic had higher detection rate (94/100, 94%) than BacT/ALERT FN Plus (80/100, 80%) (p<0.01) in the studied material. There was no significant difference in detection of anaerobic bacteria among the remaining bottle types. The 67 anaerobic bacteria that signalled positive in all four bottle types were analyzed to compare the time to detection (TTD) and isolates were directly identified by MALDI-TOF MS. There was a significant difference in TTD among the four bottle types (p<0.0001). The shortest median TTD was 18 h in BACTEC Lytic followed by BacT/ALERT FN (23.5 h), BACTEC Plus (27 h) and finally BacT/ALERT FN Plus (38 h) bottles. In contrast, MALDI-TOF MS performed similarly in all bottle types with accurate identification in 51/67 (76%) BacT/ALERT FN, 51/67 (76%) BacT/ALERT FN Plus, 53/67 (79%) BACTEC Plus and 50/67 (75%) BACTEC Lytic bottles. In conclusion, BACTEC Lytic bottles have significantly better detection rates and shorter TTD compared to the three other bottle types. The anaerobic BC bottles are equally suitable for direct MALDI-TOF MS for rapid and reliable identification of common anaerobic bacteria. Further clinical studies are warranted to investigate the performance of anaerobic BC bottles in detection of anaerobic bacteria and identification by direct MALDI-TOF MS.

The use of PCR/Electrospray Ionization-Time-of-Flight-Mass Spectrometry (PCR/ESI-TOF-MS) to detect bacterial and fungal colonization in healthy military service members.

PubMed

Vetor, Ryan; Murray, Clinton K; Mende, Katrin; Melton-Kreft, Rachel; Akers, Kevin S; Wenke, Joseph; Spirk, Tracy; Guymon, Charles; Zera, Wendy; Beckius, Miriam L; Schnaubelt, Elizabeth R; Ehrlich, Garth; Vento, Todd J

2016-07-22

The role of microbial colonization in disease is complex. Novel molecular tools to detect colonization offer theoretical improvements over traditional methods. We evaluated PCR/Electrospray Ionization-Time-of-Flight-Mass Spectrometry (PCR/ESI-TOF-MS) as a screening tool to study colonization of healthy military service members. We assessed 101 healthy Soldiers using PCR/ESI-TOF-MS on nares, oropharynx, and groin specimens for the presence of gram-positive and gram-negative bacteria (GNB), fungi, and antibiotic resistance genes. A second set of swabs was processed by traditional culture, followed by identification using the BD Phoenix automated system; comparison between PCR/ESI-TOF-MS and culture was carried out only for GNB. Using PCR/ESI-TOF-MS, at least one colonizing organism was found on each individual: mean (SD) number of organisms per subject of 11.8(2.8). The mean number of organisms in the nares, groin and oropharynx was 3.8(1.3), 3.8(1.4) and 4.2(2), respectively. The most commonly detected organisms were aerobic gram-positive bacteria: primarily coagulase-negative Staphylococcus (101 subjects: 341 organisms), Streptococcus pneumoniae (54 subjects: 57 organisms), Staphylococcus aureus (58 subjects: 80 organisms) and Nocardia asteroides (45 subjects: 50 organisms). The mecA gene was found in 96 subjects. The most commonly found GNB was Haemophilus influenzae (20 subjects: 21 organisms) and the most common anaerobe was Propionibacterium acnes (59 subjects). Saccharomyces species (30 subjects) were the most common fungi detected. Only one GNB (nares E. coli) was identified in the same subject by both diagnostic systems. PCR/ESI-TOF-MS detected common colonizing organisms and identified more typically-virulent bacteria in asymptomatic, healthy adults. PCR/ESI-TOF-MS appears to be a useful method for detecting bacterial and fungal organisms, but further clinical correlation and validation studies are needed.

An integrated microfluidic analysis microsystems with bacterial capture enrichment and in-situ impedance detection

NASA Astrophysics Data System (ADS)

Liu, Hai-Tao; Wen, Zhi-Yu; Xu, Yi; Shang, Zheng-Guo; Peng, Jin-Lan; Tian, Peng

2017-09-01

In this paper, an integrated microfluidic analysis microsystems with bacterial capture enrichment and in-situ impedance detection was purposed based on microfluidic chips dielectrophoresis technique and electrochemical impedance detection principle. The microsystems include microfluidic chip, main control module, and drive and control module, and signal detection and processing modulet and result display unit. The main control module produce the work sequence of impedance detection system parts and achieve data communication functions, the drive and control circuit generate AC signal which amplitude and frequency adjustable, and it was applied on the foodborne pathogens impedance analysis microsystems to realize the capture enrichment and impedance detection. The signal detection and processing circuit translate the current signal into impendence of bacteria, and transfer to computer, the last detection result is displayed on the computer. The experiment sample was prepared by adding Escherichia coli standard sample into chicken sample solution, and the samples were tested on the dielectrophoresis chip capture enrichment and in-situ impedance detection microsystems with micro-array electrode microfluidic chips. The experiments show that the Escherichia coli detection limit of microsystems is 5 × 104 CFU/mL and the detection time is within 6 min in the optimization of voltage detection 10 V and detection frequency 500 KHz operating conditions. The integrated microfluidic analysis microsystems laid the solid foundation for rapid real-time in-situ detection of bacteria.

[Development of molecular detection of food-borne pathogenic bacteria using miniaturized microfluidic devices].

PubMed

Iván, Kristóf; Maráz, Anna

2015-12-20

Detection and identification of food-borne pathogenic bacteria are key points for the assurance of microbiological food safety. Traditional culture-based methods are more and more replaced by or supplemented with nucleic acid based molecular techniques, targeting specific (preferably virulence) genes in the genomes. Internationally validated DNA amplification - most frequently real-time polymerase chain reaction - methods are applied by the food microbiological testing laboratories for routine analysis, which will result not only in shortening the time for results but they also improve the performance characteristics (e.g. sensitivity, specificity) of the methods. Beside numerous advantages of the polymerase chain reaction based techniques for routine microbiological analysis certain drawbacks have to be mentioned, such as the high cost of the equipment and reagents, as well as the risk of contamination of the laboratory environment by the polymerase chain reaction amplicons, which require construction of an isolated laboratory system. Lab-on-a-chip systems can integrate most of these laboratory processes within a miniaturized device that delivers the same specificity and reliability as the standard protocols. The benefits of miniaturized devices are: simple - often automated - use, small overall size, portability, sterility due to single use possibility. These miniaturized rapid diagnostic tests are being researched and developed at the best research centers around the globe implementing various sample preparation and molecular DNA amplification methods on-chip. In parallel, the aim of the authors' research is to develop microfluidic Lab-on-a-chip devices for the detection and identification of food-borne pathogenic bacteria.

Detection of microbial communities in continuous and discontinuous membrane bioreactor using high-density oligonucleotide Microarray

NASA Astrophysics Data System (ADS)

Duan, Liang; Song, Yonghui; Xia, Siqing; Hermanowicz, Slawomir W.

2010-11-01

This study compared the whole composition of microbial communities in continuous-flow (MBR) and batch-fed (discontinuous) (MSBR) aerobic membrane bioreactors using high-density universal 16S rRNA Microarray. The array includes 506,944 probes targeted to 8935 clusters in 16S rRNA gene sequences. The Microarray results showed that both MBR and MSBR had high microbial diversity. 1126 and 1002 bacterial subfamilies were detected and can separate as 37 and 32 phyla in MBR and MSBR, respectively. Proteobacteria was the predominant phylum, 703 and 597 subfamilies were found in two systems, which constituted 62.4% and 59.6% of the whole bacteria. Gamma- and Alpha-were the dominant classes in Proteobacteria. It occupied 38.1% and 26.3%, 31.2% and 39.2% for MBR and MSBR, respectively. Bacteroidetes, Firmicutes and Actinobacteria were the subdominant groups, occupying around 9.4% and 7.6%, 6.1% and 6.5%, 6.0% and 9.0% of the total bacteria in two reactors. Some bacterial groups such as Acidobacteria, Chloroflexi, Cyanobacteria, Verrucomicrobia and Spirochaetes also found more than 15 subfamilies. All the results indicated that the MBR system had more bacteria community diversity than MSBR's. Moreover, it was very interested that MBR and MSBR had almost the same bacterial composition except Enterobacteriaceae. 63 OTUs of Enterobacteriaceae were detected in MBR, while just 10 OTUs were found in MSBR. That's one of the reasons leading to the difference of the bacterial diversity between two bioreactors.

Highly labeled methylene blue-ds DNA silica nanoparticles for signal enhancement of immunoassays: application to the sensitive detection of bacteria in human platelet concentrates.

PubMed

Bonnet, Romaric; Farre, Carole; Valera, Lionel; Vossier, Ludivine; Léon, Fanny; Dagland, Typhaine; Pouzet, Agnès; Jaffrézic-Renault, Nicole; Fareh, Jeannette; Fournier-Wirth, Chantal; Chaix, Carole

2018-05-15

A nanoparticle-based electrochemical sandwich immunoassay was developed for bacteria detection in platelet concentrates. For the assay, magnetic beads were functionalized with antibodies to allow the specific capture of bacteria from the complex matrix, and innovative methylene blue-DNA/nanoparticle assemblies provided the electrochemical response for amplified detection. This nanoparticular system was designed as a temperature-sensitive nano-tool for electrochemical detection. First, oligonucleotide-functionalized nanoparticles were obtained by direct synthesis of the DNA strands on the nanoparticle surface using an automated oligonucleotide synthesizer. Densely packed DNA coverage was thus obtained. Then, DNA duplexes were constructed on the NP surface with a complementary strand bearing a 3 methylene blue tag. This strategy ultimately produced highly functionalized nanoparticles with electrochemical markers. These assemblies enabled amplification of the electrochemical signal, resulting in a very good sensitivity. A proof-of-concept was carried out for E. coli detection in human platelet concentrates. Bacterial contamination of this complex biological matrix is the highest residual infectious risk in blood transfusion. The development of a rapid assay that could reach 10-102 CFU mL-1 sensitivity is a great challenge. The nanoparticle-based electrochemical sandwich immunoassay carried out on a boron doped diamond electrode proved to be sensitive for E. coli detection in human platelets. Two antibody pairs were used to develop either a generic assay against certain Gram negative strains or a specific assay for E. coli. The methylene blue-DNA/nanoparticles amplify sensitivity ×1000 compared with the assay run without NPs for electrochemical detection. A limit of detection of 10 CFU mL-1 in a biological matrix was achieved for E. coli using the highly specific antibody pair.

Microbial contamination of vegetable crop and soil profile in arid regions under controlled application of domestic wastewater

PubMed Central

Balkhair, Khaled S.

2015-01-01

Increasing lack of potable water in arid countries leads to the use of treated wastewater for crop production. However, the use of inappropriate irrigation practices could result in a serious contamination risk to plants, soils, and groundwater with sewage water. This research was initiated in view to the increasing danger of vegetable crops and groundwater contamination with pathogenic bacteria due to wastewater land application. The research was designed to study: (1) the effect of treated wastewater irrigation on the yield and microbial contamination of the radish plant under field conditions; (2) contamination of the agricultural soil profile with fecal coliform bacteria. Effluent from a domestic wastewater treatment plant (100%) in Jeddah city, Saudi Arabia, was diluted to 80% and 40% with the groundwater of the experimental site constituting three different water qualities plus groundwater as control. Radish plant was grown in two consecutive seasons under two drip irrigation systems and four irrigation water qualities. Upon harvesting, plant weight per ha, total bacterial, fecal coliform, fecal streptococci were detected per 100 g of dry matter and compared with the control. The soil profile was also sampled at an equal distance of 3 cm from soil surface for fecal coliform detection. The results indicated that the yield increased significantly under the subsurface irrigation system and the control water quality compared to surface irrigation system and other water qualities. There was a considerable drop in the count of all bacteria species under the subsurface irrigation system compared to surface irrigation. The bacterial count/g of the plant shoot system increased as the percentage of wastewater in the irrigation water increased. Most of the fecal coliform bacteria were deposited in the first few centimeters below the column inlet and the profile exponentially decreased with increasing depth. PMID:26858571

Transmutation of Personal Glucose Meters into Portable and Highly Sensitive Microbial Pathogen Detection Platform.

PubMed

Wang, Zhenzhen; Chen, Zhaowei; Gao, Nan; Ren, Jinsong; Qu, Xiaogang

2015-10-07

Herein, for the first time, we presented a simple and general approach by using personal glucose meters (PGM) for portable and ultrasensitive detection of microbial pathogens. Upon addition of pathogenic bacteria, glucoamylase-quaternized magnetic nanoparticles (GA-QMNPS) conjugates were disrupted by the competitive multivalent interactions between bacteria and QMNPS, resulting in the release of GA. After magnetic separation, the free GA could catalyze the hydrolysis of amylose into glucose for quantitative readout by PGM. In such way, PGM was transmuted into a bacterial detection device and extremely low detection limits down to 20 cells mL(-1) was achieved. More importantly, QMNPS could inhibit the growth of the bacteria and destroy its cellular structure, which enabled bacteria detection and inhibition simultaneously. The simplicity, portability, sensitivity and low cost of presented work make it attractive for clinical applications. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Enzyme Characteristics of β-d-Galactosidase- and β-d-Glucuronidase-Positive Bacteria and Their Interference in Rapid Methods for Detection of Waterborne Coliforms and Escherichia coli

PubMed Central

Tryland, I.; Fiksdal, L.

1998-01-01

Bacteria which were β-d-galactosidase and β-d-glucuronidase positive or expressed only one of these enzymes were isolated from environmental water samples. The enzymatic activity of these bacteria was measured in 25-min assays by using the fluorogenic substrates 4-methylumbelliferyl-β-d-galactoside and 4-methylumbelliferyl-β-d-glucuronide. The enzyme activity, enzyme induction, and enzyme temperature characteristics of target and nontarget bacteria in assays aimed at detecting coliform bacteria and Escherichia coli were investigated. The potential interference of false-positive bacteria was evaluated. Several of the β-d-galactosidase-positive nontarget bacteria but none of the β-d-glucuronidase-positive nontarget bacteria contained unstable enzyme at 44.5°C. The activity of target bacteria was highly inducible. Nontarget bacteria were induced much less or were not induced by the inducers used. The results revealed large variations in the enzyme levels of different β-d-galactosidase- and β-d-glucuronidase-positive bacteria. The induced and noninduced β-d-glucuronidase activities of Bacillus spp. and Aerococcus viridans were approximately the same as the activities of induced E. coli. Except for some isolates identified as Aeromonas spp., all of the induced and noninduced β-d-galactosidase-positive, noncoliform isolates exhibited at least 2 log units less mean β-d-galactosidase activity than induced E. coli. The noncoliform bacteria must be present in correspondingly higher concentrations than those of target bacteria to interfere in the rapid assay for detection of coliform bacteria. PMID:9501441

The use of a differential fluorescent staining method to detect bacteriuria.

PubMed

Ciancaglini, Ettore; Fazii, Paolo; Sforza, Giuseppe Riario

2004-01-01

This report describes a differential staining method which distinguishes gram-positive from gram-negative bacteria in fluorescence. Gram-positive bacteria appear yellow and gram-negative bacteria appear green. The method is based on two fluorochromes, one acting in the wavelength of red, i.e. the acridine orange, and another acting in the wavelength of green, i.e. the fluorescein, which together form a red/ green system. In this report we compared the accuracy of the differential fluorescent staining method and the Gram stain in screening for bacteriuria, as detected by conventional cultures. A total of 1487 urine samples were tested. 289 cultures were positive. 237 specimens grew a single organism at 10(5) and 10(4) CFU/ml. 224 smears were detected by the differential fluorescent staining method and 162 were detected by Gram stain. 1198 samples failed to grow organisms at 10(5) and 10(4) CFU/ml. 107 smears were falsely positive by the fluorescent staining procedure and 289 were falsely positive by the Gram stain. On the basis of the culture results, the sensitivity of the differential fluorescent staining method was 94.5% and that of the Gram stain 68.3%. The specificity of the fluorescent staining procedure was 91.6% and that of the Gram stain 75.8%. The positive predictive value and the negative predictive value of the fluorescent staining method were 67.6% and 98.8%, respectively. Those of the Gram stain were 35.9% and 92.3%, respectively. A wide range of microbiological and chemical techniques are available to identify bacteria in urine. This fluorescent staining method represents a simple, rapid, reliable method with low-running costs. The main advantage of this technique is that it enables the microbiologist to exclude the presence of bacteria in the urine within a short time after specimen receipt and to eliminate a large number of specimens for culture with significant cost saving. Another advantage of the method is that it allows to distinguish gram-positive from gram-negative bacteria in positive slides on the same day the sample is obtained. The stained smears were easily interpreted, even when the bacterial counts in the specimen were low.

Computational determination of the effects of virulent Escherichia coli and salmonella bacteriophages on human gut.

PubMed

Mostafa, Marwa Mostafa; Nassef, Mohammad; Badr, Amr

2016-10-01

Salmonella and Escherichia coli are different types of bacteria that cause food poisoning in humans. In the elderly, infants and people with chronic conditions, it is very dangerous if Salmonella or E. coli gets into the bloodstream and then they must be treated by phage therapy. Treating Salmonella and E. coli by phage therapy affects the gut flora. This research paper presents a system for detecting the effects of virulent E. coli and Salmonella bacteriophages on human gut. A method based on Domain-Domain Interactions (DDIs) model is implemented in the proposed system to determine the interactions between the proteins of human gut bacteria and the proteins of bacteriophages that infect virulent E. coli and Salmonella. The system helps gastroenterologists to realize the effect of injecting bacteriophages that infect virulent E. coli and Salmonella on the human gut. By testing the system over Enterobacteria phage 933W, Enterobacteria phage VT2-Sa and Enterobacteria phage P22, it resulted in four interactions between the proteins of the bacteriophages that infect E. coli O157:H7, E. coli O104:H4 and Salmonella typhimurium and the proteins of human gut bacterium strains. Several effects were detected such as: antibacterial activity against a number of bacterial species in human gut, regulation of cellular differentiation and organogenesis during gut, lung, and heart development, ammonia assimilation in bacteria, yeasts, and plants, energizing defense system and its function in the detoxification of lipopolysaccharide, and in the prevention of bacterial translocation in human gut. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

Genus-wide comparison of Pseudovibrio bacterial genomes reveal diverse adaptations to different marine invertebrate hosts.

PubMed

Alex, Anoop; Antunes, Agostinho

2018-01-01

Bacteria belonging to the genus Pseudovibrio have been frequently found in association with a wide variety of marine eukaryotic invertebrate hosts, indicative of their versatile and symbiotic lifestyle. A recent comparison of the sponge-associated Pseudovibrio genomes has shed light on the mechanisms influencing a successful symbiotic association with sponges. In contrast, the genomic architecture of Pseudovibrio bacteria associated with other marine hosts has received less attention. Here, we performed genus-wide comparative analyses of 18 Pseudovibrio isolated from sponges, coral, tunicates, flatworm, and seawater. The analyses revealed a certain degree of commonality among the majority of sponge- and coral-associated bacteria. Isolates from other marine invertebrate host, tunicates, exhibited a genetic repertoire for cold adaptation and specific metabolic abilities including mucin degradation in the Antarctic tunicate-associated bacterium Pseudovibrio sp. Tun.PHSC04_5.I4. Reductive genome evolution was simultaneously detected in the flatworm-associated bacteria and the sponge-associated bacterium P. axinellae AD2, through the loss of major secretion systems (type III/VI) and virulence/symbioses factors such as proteins involved in adhesion and attachment to the host. Our study also unraveled the presence of a CRISPR-Cas system in P. stylochi UST20140214-052 a flatworm-associated bacterium possibly suggesting the role of CRISPR-based adaptive immune system against the invading virus particles. Detection of mobile elements and genomic islands (GIs) in all bacterial members highlighted the role of horizontal gene transfer for the acquisition of novel genetic features, likely enhancing the bacterial ecological fitness. These findings are insightful to understand the role of genome diversity in Pseudovibrio as an evolutionary strategy to increase their colonizing success across a wide range of marine eukaryotic hosts.

Genus-wide comparison of Pseudovibrio bacterial genomes reveal diverse adaptations to different marine invertebrate hosts

PubMed Central

Alex, Anoop

2018-01-01

Bacteria belonging to the genus Pseudovibrio have been frequently found in association with a wide variety of marine eukaryotic invertebrate hosts, indicative of their versatile and symbiotic lifestyle. A recent comparison of the sponge-associated Pseudovibrio genomes has shed light on the mechanisms influencing a successful symbiotic association with sponges. In contrast, the genomic architecture of Pseudovibrio bacteria associated with other marine hosts has received less attention. Here, we performed genus-wide comparative analyses of 18 Pseudovibrio isolated from sponges, coral, tunicates, flatworm, and seawater. The analyses revealed a certain degree of commonality among the majority of sponge- and coral-associated bacteria. Isolates from other marine invertebrate host, tunicates, exhibited a genetic repertoire for cold adaptation and specific metabolic abilities including mucin degradation in the Antarctic tunicate-associated bacterium Pseudovibrio sp. Tun.PHSC04_5.I4. Reductive genome evolution was simultaneously detected in the flatworm-associated bacteria and the sponge-associated bacterium P. axinellae AD2, through the loss of major secretion systems (type III/VI) and virulence/symbioses factors such as proteins involved in adhesion and attachment to the host. Our study also unraveled the presence of a CRISPR-Cas system in P. stylochi UST20140214-052 a flatworm-associated bacterium possibly suggesting the role of CRISPR-based adaptive immune system against the invading virus particles. Detection of mobile elements and genomic islands (GIs) in all bacterial members highlighted the role of horizontal gene transfer for the acquisition of novel genetic features, likely enhancing the bacterial ecological fitness. These findings are insightful to understand the role of genome diversity in Pseudovibrio as an evolutionary strategy to increase their colonizing success across a wide range of marine eukaryotic hosts. PMID:29775460

133Xe contamination found in internal bacteria filter of xenon ventilation system.

PubMed

Hackett, Michael T; Collins, Judith A; Wierzbinski, Rebecca S

2003-09-01

We report on (133)Xe contamination found in the reusable internal bacteria filter of our xenon ventilation system. Internal bacteria filters (n = 6) were evaluated after approximately 1 mo of normal use. The ventilation system was evacuated twice to eliminate (133)Xe in the system before removal of the filter. Upon removal, the filter was monitored using a survey meter with an energy-compensated probe and was imaged on a scintillation camera. The filter was monitored and imaged over several days and was stored in a fume hood. Estimated (133)Xe activity in each filter immediately after removal ranged from 132 to 2,035 kBq (3.6-55.0 micro Ci), based on imaging. Initial surface radiation levels ranged from 0.4 to 4.5 micro Sv/h (0.04-0.45 mrem/h). The (133)Xe activity did not readily leave the filter over time (i.e., time to reach half the counts of the initial decay-corrected image ranged from <6 to >72 h). The majority of the image counts (approximately 70%) were seen in 2 distinctive areas in the filter. They corresponded to sites where the manufacturer used polyurethane adhesive to attach the fiberglass filter medium to the filter housing. (133)Xe contamination within the reusable internal bacteria filter of our ventilation system was easily detected by a survey meter and imaging. Although initial activities and surface radiation levels were low, radiation safety practices would dictate that a (133)Xe-contaminated bacteria filter be stored preferably in a fume hood until it cannot be distinguished from background before autoclaving or disposal.

[Investigation of bacterial and viral etiology in community acquired central nervous system infections with molecular methods].

PubMed

Kahraman, Hasip; Tünger, Alper; Şenol, Şebnem; Gazi, Hörü; Avcı, Meltem; Örmen, Bahar; Türker, Nesrin; Atalay, Sabri; Köse, Şükran; Ulusoy, Sercan; Işıkgöz Taşbakan, Meltem; Sipahi, Oğuz Reşat; Yamazhan, Tansu; Gülay, Zeynep; Alp Çavuş, Sema; Pullukçu, Hüsnü

2017-07-01

In this multicenter prospective cohort study, it was aimed to evaluate the bacterial and viral etiology in community-acquired central nervous system infections by standart bacteriological culture and multiplex polymerase chain reaction (PCR) methods. Patients hospitalized with central nervous system infections between April 2012 and February 2014 were enrolled in the study. Demographic and clinical information of the patients were collected prospectively. Cerebrospinal fluid (CSF) samples of the patients were examined by standart bacteriological culture methods, bacterial multiplex PCR (Seeplex meningitis-B ACE Detection (Streptococcus pneumoniae, Neisseria meningitidis, Haemophilus influenzae, Listeria monocytogenes, Group B streptococci) and viral multiplex PCR (Seeplex meningitis-V1 ACE Detection kits herpes simplex virus-1 (HSV1), herpes simplex virus-2 (HSV2), varicella zoster virus (VZV), cytomegalovirus (CMV), Epstein Barr virus (EBV) and human herpes virus 6 (HHV6)) (Seeplex meningitis-V2 ACE Detection kit (enteroviruses)). Patients were classified as purulent meningitis, aseptic meningitis and encephalitis according to their clinical, CSF (leukocyte level, predominant cell type, protein and glucose (blood/CSF) levels) and cranial imaging results. Patients who were infected with a pathogen other than the detection of the kit or diagnosed as chronic meningitis and other diseases during the follow up, were excluded from the study. A total of 79 patients (28 female, 51 male, aged 42.1 ± 18.5) fulfilled the study inclusion criteria. A total of 46 patients were classified in purulent meningitis group whereas 33 were in aseptic meningitis/encephalitis group. Pathogens were detected by multiplex PCR in 41 patients. CSF cultures were positive in 10 (21.7%) patients (nine S.pneumoniae, one H.influenzae) and PCR were positive for 27 (58.6%) patients in purulent meningitis group. In this group one type of bacteria were detected in 18 patients (14 S.pneumoniae, two N.meningitidis, one H.influenzae, one L.monocytogenes). Besides, it is noteworthy that multiple pathogens were detected such as bacteria-virus combination in eight patients and two different bacteria in one patient. In the aseptic meningitis/encephalitis group, pathogens were detected in 14 out of 33 patients; single type of viruses in 11 patients (seven enterovirus, two HSV1, one HSV2, one VZV) and two different viruses were determined in three patients. These data suggest that multiplex PCR methods may increase the isolation rate of pathogens in central nervous system infections. Existence of mixed pathogen growth is remarkable in our study. Further studies are needed for the clinical relevance of this result.

DETECTION,QUANTIFICATION AND CHARACTERIZATION OF MYCOBACTERIUM AVIUM COMPLEX (MAC) ORGANISMS IN DRINKING WATER.

EPA Science Inventory

Bacteria belonging to the Mycobacterium avium complex (MAC), including Mycobacterium avium and M. intracellulare, are clinically relevant and cause a myriad of opportunistic infections. Children, the elderly, and persons with previous lung conditions or immune system dysfunction...

Evaluation of Polymerase Chain Reaction for Detecting Coliform Bacteria in Drinking Water Sources.

PubMed

Isfahani, Bahram Nasr; Fazeli, Hossein; Babaie, Zeinab; Poursina, Farkhondeh; Moghim, Sharareh; Rouzbahani, Meisam

2017-01-01

Coliform bacteria are used as indicator organisms for detecting fecal pollution in water. Traditional methods including microbial culture tests in lactose-containing media and enzyme-based tests for the detection of β-galactosidase; however, these methods are time-consuming and less specific. The aim of this study was to evaluate polymerase chain reaction (PCR) for detecting coliform. Totally, 100 of water samples from Isfahan drinking water source were collected. Coliform bacteria and Escherichia coli were detected in drinking water using LacZ and LamB genes in PCR method performed in comparison with biochemical tests for all samples. Using phenotyping, 80 coliform isolates were found. The results of the biochemical tests illustrated 78.7% coliform bacteria and 21.2% E. coli . PCR results for LacZ and LamB genes were 67.5% and 17.5%, respectively. The PCR method was shown to be an effective, sensitive, and rapid method for detecting coliform and E. coli in drinking water from the Isfahan drinking water sources.

Microbial Load Monitor

NASA Technical Reports Server (NTRS)

Gibson, S. F.; Royer, E. R.

1979-01-01

The Microbial Load Monitor (MLM) is an automated and computerized system for detection and identification of microorganisms. Additionally, the system is designed to enumerate and provide antimicrobic susceptibility profiles for medically significant bacteria. The system is designed to accomplish these tasks in a time of 13 hours or less versus the traditional time of 24 hours for negatives and 72 hours or more for positives usually required for standard microbiological analysis. The MLM concept differs from other methods of microbial detection in that the system is designed to accept raw untreated clinical samples and to selectively identify each group or species that may be present in a polymicrobic sample.

Does virus-bacteria coinfection increase the clinical severity of acute respiratory infection?

PubMed

Damasio, Guilherme A C; Pereira, Luciane A; Moreira, Suzana D R; Duarte dos Santos, Claudia N; Dalla-Costa, Libera M; Raboni, Sonia M

2015-09-01

This retrospective cohort study investigated the presence of bacteria in respiratory secretions of patients hospitalized with acute respiratory infections and analyzed the impact of viral and bacterial coinfection on severity and the mortality rate. A total of 169 patients with acute respiratory infections were included, viruses and bacteria in respiratory samples were detected using molecular methods. Among all samples, 73.3% and 59.7% were positive for viruses and bacteria, respectively; 45% contained both virus and bacteria. Bacterial coinfection was more frequent in patients infected by community respiratory viruses than influenza A H1N1pdm (83.3% vs. 40.6%). The most frequently bacteria detected were Streptococcus pneumoniae and Haemophilus influenzae. Both species were co-detected in 54 patients and identified alone in 22 and 21 patients, respectively. Overall, there were no significant differences in the period of hospitalization, severity, or mortality rate between patients infected with respiratory viruses alone and those coinfected by viruses and bacteria. The detection of mixed respiratory pathogens is frequent in hospitalized patients with acute respiratory infections, but its impact on the clinical outcome does not appear substantial. However, it should be noted that most of the patients received broad-spectrum antibiotic therapy, which may have contributed to this favorable outcome. © 2015 Wiley Periodicals, Inc.

Quantification and risks associated with bacterial aerosols near domestic greywater-treatment systems.

PubMed

Benami, Maya; Busgang, Allison; Gillor, Osnat; Gross, Amit

2016-08-15

Greywater (GW) reuse can alleviate water stress by lowering freshwater consumption. However, GW contains pathogens that may compromise public health. During the GW-treatment process, bioaerosols can be produced and may be hazardous to human health if inhaled, ingested, or come in contact with skin. Using air-particle monitoring, BioSampler®, and settle plates we sampled bioaerosols emitted from recirculating vertical flow constructed wetlands (RVFCW) - a domestic GW-treatment system. An array of pathogens and indicators were monitored using settle plates and by culturing the BioSampler® liquid. Further enumeration of viable pathogens in the BioSampler® liquid utilized a newer method combining the benefits of enrichment with molecular detection (MPN-qPCR). Additionally, quantitative microbial risk assessment (QMRA) was applied to assess risks of infection from a representative skin pathogen, Staphylococcus aureus. According to the settle-plate technique, low amounts (0-9.7×10(4)CFUm(-2)h(-1)) of heterotrophic bacteria, Staphylococcus spp., Pseudomonas spp., Klebsiella pneumoniae, Enterococcus spp., and Escherichia coli were found to aerosolize up to 1m away from the GW systems. At the 5m distance amounts of these bacteria were not statistically different (p>0.05) from background concentrations tested over 50m away from the systems. Using the BioSampler®, no bacteria were detected before enrichment of the GW-aerosols. However, after enrichment, using an MPN-qPCR technique, viable indicators and pathogens were occasionally detected. Consequently, the QMRA results were below the critical disability-adjusted life year (DALY) safety limits, a measure of overall disease burden, for S. aureus under the tested exposure scenarios. Our study suggests that health risks from aerosolizing pathogens near RVFCW GW-treatment systems are likely low. This study also emphasizes the growing need for standardization of bioaerosol-evaluation techniques to provide more accurate quantification of small amounts of viable, aerosolized bacterial pathogens. Copyright © 2016 Elsevier B.V. All rights reserved.

Antibody purification-independent microarrays (PIM) by direct bacteria spotting on TiO2-treated slides.

PubMed

De Marni, Marzia L; Monegal, Ana; Venturini, Samuele; Vinati, Simone; Carbone, Roberta; de Marco, Ario

2012-02-01

The preparation of effective conventional antibody microarrays depends on the availability of high quality material and on the correct accessibility of the antibody active moieties following their immobilization on the support slide. We show that spotting bacteria that expose recombinant antibodies on their external surface directly on nanostructured-TiO(2) or epoxy slides (purification-independent microarray - PIM) is a simple and reliable alternative for preparing sensitive and specific microarrays for antigen detection. Variable domains of single heavy-chain antibodies (VHHs) against fibroblast growth factor receptor 1 (FGFR1) were used to capture the antigen diluted in serum or BSA solution. The FGFR1 detection was performed by either direct antigen labeling or using a sandwich system in which FGFR1 was first bound to its antibody and successively identified using a labeled FGF. In both cases the signal distribution within each spot was uniform and spot morphology regular. The signal-to-noise ratio of the signal was extremely elevated and the specificity of the system was proved statistically. The LOD of the system for the antigen was calculated being 0.4ng/mL and the dynamic range between 0.4ng/mL and 10μg/mL. The microarrays prepared with bacteria exposing antibodies remain fully functional for at least 31 days after spotting. We finally demonstrated that the method is suitable for other antigen-antibody pairs and expect that it could be easily adapted to further applications such as the display of scFv and IgG antibodies or the autoantibody detection using protein PIMs. Copyright © 2011. Published by Elsevier Inc.

Raman microspectrometer combined with scattering microscopy and lensless imaging for bacteria identification

NASA Astrophysics Data System (ADS)

Strola, S. A.; Schultz, E.; Allier, C. P.; DesRoches, B.; Lemmonier, J.; Dinten, J.-M.

2013-03-01

In this paper, we report on a compact prototype capable both of lensfree imaging, Raman spectrometry and scattering microscopy from bacteria samples. This instrument allows high-throughput real-time characterization without the need of markers, making it potentially suitable to field label-free biomedical and environmental applications. Samples are illuminated from above with a focused-collimated 532nm laser beam and can be x-y-z scanned. The bacteria detection is based on emerging lensfree imaging technology able to localize cells of interest over a large field-of-view of 24mm2. Raman signal and scattered light are then collected by separate measurement arms simultaneously. In the first arm the emission light is fed by a fiber into a prototype spectrometer, developed by Tornado Spectral System based on Tornado's High Throughput Virtual Slit (HTVS) novel technology. The enhanced light throughput in the spectral region of interest (500-1800 cm-1) reduces Raman acquisition time down to few seconds, thus facilitating experimental protocols and avoiding the bacteria deterioration induced by laser thermal heating. Scattered light impinging in the second arm is collected onto a charge-coupled-device. The reconstructed image allows studying the single bacteria diffraction pattern and their specific structural features. The characterization and identification of different bacteria have been performed to validate and optimize the acquisition system and the component setup. The results obtained demonstrate the benefits of these three techniques combination by providing the precise bacteria localization, their chemical composition and a morphology description. The procedure for a rapid identification of particular pathogen bacteria in a sample is illustrated.

Sensitive, Rapid Detection of Bacterial Spores

NASA Technical Reports Server (NTRS)

Kern, Roger G.; Venkateswaran, Kasthuri; Chen, Fei; Pickett, Molly; Matsuyama, Asahi

2009-01-01

A method of sensitive detection of bacterial spores within delays of no more than a few hours has been developed to provide an alternative to a prior three-day NASA standard culture-based assay. A capability for relatively rapid detection of bacterial spores would be beneficial for many endeavors, a few examples being agriculture, medicine, public health, defense against biowarfare, water supply, sanitation, hygiene, and the food-packaging and medical-equipment industries. The method involves the use of a commercial rapid microbial detection system (RMDS) that utilizes a combination of membrane filtration, adenosine triphosphate (ATP) bioluminescence chemistry, and analysis of luminescence images detected by a charge-coupled-device camera. This RMDS has been demonstrated to be highly sensitive in enumerating microbes (it can detect as little as one colony-forming unit per sample) and has been found to yield data in excellent correlation with those of culture-based methods. What makes the present method necessary is that the specific RMDS and the original protocols for its use are not designed for discriminating between bacterial spores and other microbes. In this method, a heat-shock procedure is added prior to an incubation procedure that is specified in the original RMDS protocols. In this heat-shock procedure (which was also described in a prior NASA Tech Briefs article on enumerating sporeforming bacteria), a sample is exposed to a temperature of 80 C for 15 minutes. Spores can survive the heat shock, but nonspore- forming bacteria and spore-forming bacteria that are not in spore form cannot survive. Therefore, any colonies that grow during incubation after the heat shock are deemed to have originated as spores.

Comparative quantification of human intestinal bacteria based on cPCR and LDR/LCR

PubMed Central

Tang, Zhou-Rui; Li, Kai; Zhou, Yu-Xun; Xiao, Zhen-Xian; Xiao, Jun-Hua; Huang, Rui; Gu, Guo-Hao

2012-01-01

AIM: To establish a multiple detection method based on comparative polymerase chain reaction (cPCR) and ligase detection reaction (LDR)/ligase chain reaction (LCR) to quantify the intestinal bacterial components. METHODS: Comparative quantification of 16S rDNAs from different intestinal bacterial components was used to quantify multiple intestinal bacteria. The 16S rDNAs of different bacteria were amplified simultaneously by cPCR. The LDR/LCR was examined to actualize the genotyping and quantification. Two beneficial (Bifidobacterium, Lactobacillus) and three conditionally pathogenic bacteria (Enterococcus, Enterobacterium and Eubacterium) were used in this detection. With cloned standard bacterial 16S rDNAs, standard curves were prepared to validate the quantitative relations between the ratio of original concentrations of two templates and the ratio of the fluorescence signals of their final ligation products. The internal controls were added to monitor the whole detection flow. The quantity ratio between two bacteria was tested. RESULTS: cPCR and LDR revealed obvious linear correlations with standard DNAs, but cPCR and LCR did not. In the sample test, the distributions of the quantity ratio between each two bacterial species were obtained. There were significant differences among these distributions in the total samples. But these distributions of quantity ratio of each two bacteria remained stable among groups divided by age or sex. CONCLUSION: The detection method in this study can be used to conduct multiple intestinal bacteria genotyping and quantification, and to monitor the human intestinal health status as well. PMID:22294830

Comparative quantification of human intestinal bacteria based on cPCR and LDR/LCR.

PubMed

Tang, Zhou-Rui; Li, Kai; Zhou, Yu-Xun; Xiao, Zhen-Xian; Xiao, Jun-Hua; Huang, Rui; Gu, Guo-Hao

2012-01-21

To establish a multiple detection method based on comparative polymerase chain reaction (cPCR) and ligase detection reaction (LDR)/ligase chain reaction (LCR) to quantify the intestinal bacterial components. Comparative quantification of 16S rDNAs from different intestinal bacterial components was used to quantify multiple intestinal bacteria. The 16S rDNAs of different bacteria were amplified simultaneously by cPCR. The LDR/LCR was examined to actualize the genotyping and quantification. Two beneficial (Bifidobacterium, Lactobacillus) and three conditionally pathogenic bacteria (Enterococcus, Enterobacterium and Eubacterium) were used in this detection. With cloned standard bacterial 16S rDNAs, standard curves were prepared to validate the quantitative relations between the ratio of original concentrations of two templates and the ratio of the fluorescence signals of their final ligation products. The internal controls were added to monitor the whole detection flow. The quantity ratio between two bacteria was tested. cPCR and LDR revealed obvious linear correlations with standard DNAs, but cPCR and LCR did not. In the sample test, the distributions of the quantity ratio between each two bacterial species were obtained. There were significant differences among these distributions in the total samples. But these distributions of quantity ratio of each two bacteria remained stable among groups divided by age or sex. The detection method in this study can be used to conduct multiple intestinal bacteria genotyping and quantification, and to monitor the human intestinal health status as well.

Nonenzymatic microorganism identification based on ribosomal RNA

NASA Astrophysics Data System (ADS)

Ives, Jeffrey T.; Pierini, Alicia M.; Stokes, Jeffrey A.; Wahlund, Thomas M.; Read, Betsy; Bechtel, James H.; Bronk, Burt V.

1999-11-01

Effective defense against biological warfare (BW) agents requires rapid, fieldable and accurate systems. For micro- organisms like bacteria and viruses, ribosomal RNA (rRNA) provides a valuable target with multiple advantages of species specificity and intrinsic target amplification. Vegetative and spore forms of bacteria contain approximately 104 copies of rRNA. Direct detection of rRNA copies can eliminate some of the interference and preparation difficulties involved in enzymatic amplification methods. In order to apply the advantages of rRNA to BW defense, we are developing a fieldable system based on 16S rRNA, physical disruption of the micro-organism, solid phase hybridization, and fluorescence detection. Our goals include species-specific identification, complete operation from raw sample to identification in 15 minutes or less, and compact, fieldable instrumentation. Initial work on this project has investigated the lysis and hybridization steps, the species-specificity of oligonucleotides probes, and the development of a novel electromagnetic method to physically disrupt the micro- organisms. Target bacteria have been Escherichia coli (E. coli) and Bacillus subtilis (B. subtilis). Continuing work includes further development of methods to rapidly disrupt the micro-organisms and release the rRNA, improved integration and processing, and extension to bacterial and mammalian viruses like MS2 and vesicular stomatitis virus.

Eavesdropping and crosstalk between secreted quorum sensing peptide signals that regulate bacteriocin production in Streptococcus pneumoniae.

PubMed

Miller, Eric L; Kjos, Morten; Abrudan, Monica I; Roberts, Ian S; Veening, Jan-Willem; Rozen, Daniel E

2018-06-13

Quorum sensing (QS), where bacteria secrete and respond to chemical signals to coordinate population-wide behaviors, has revealed that bacteria are highly social. Here, we investigate how diversity in QS signals and receptors can modify social interactions controlled by the QS system regulating bacteriocin secretion in Streptococcus pneumoniae, encoded by the blp operon (bacteriocin-like peptide). Analysis of 4096 pneumococcal genomes detected nine blp QS signals (BlpC) and five QS receptor groups (BlpH). Imperfect concordance between signals and receptors suggested widespread social interactions between cells, specifically eavesdropping (where cells respond to signals that they do not produce) and crosstalk (where cells produce signals that non-clones detect). This was confirmed in vitro by measuring the response of reporter strains containing six different blp QS receptors to cognate and non-cognate peptides. Assays between pneumococcal colonies grown adjacent to one another provided further evidence that crosstalk and eavesdropping occur at endogenous levels of signal secretion. Finally, simulations of QS strains producing bacteriocins revealed that eavesdropping can be evolutionarily beneficial even when the affinity for non-cognate signals is very weak. Our results highlight that social interactions can mediate intraspecific competition among bacteria and reveal that competitive interactions can be modified by polymorphic QS systems.

Differential fluorescent staining method for detection of bacteria in blood cultures, cerebrospinal fluid and other clinical specimens.

PubMed

Fazii, P; Ciancaglini, E; Riario Sforza, G

2002-05-01

The aim of this study was to evaluate a differential staining method to distinguish gram-positive from gram-negative bacteria in fluorescence. The method is based on two fluorochromes, one acting in the wavelength of red, i.e. the acridine orange, and another acting in the wavelength of green, i.e. the fluorescein. With this method, gram-positive bacteria appear yellow and gram-negative bacteria appear green. In view of the importance of a rapid aetiological diagnosis in cases of septicaemia, the differential staining method in fluorescence was compared with Gram stain for the detection of bacteria in blood. Of 5,820 blood cultures entered into the study and identified by the Bactec 9120 fluorescent series instrument (Becton Dickinson Europe, France), 774 were positive. Of the 774 positive cultures, 689 yielded only a single organism. The differential staining method in fluorescence detected 626 of the 689 cultures, while Gram stain detected 468. On the basis of these results, the sensitivity of the differential staining method in fluorescence was 90.9%, while that of Gram stain was 67.9%. The difference between the two methods was statistically significant ( P<0.001). The differential fluorescent staining method was more sensitive than Gram stain in the detection of bacteria in blood cultures during the incubation period. This technique provides a rapid, simple and highly sensitive staining method that can be used in conjunction with subculture methods. Whereas subculture requires an incubation period of 18-24 h, the fluorescent staining technique can detect bacteria on the same day that smears are prepared and examined. The differential fluorescent staining method was also evaluated for its ability to detect microorganisms in cerebrospinal fluid and other clinical specimens. The microorganisms were easily detected, even when bacterial counts in the specimens were low.

Eukaryote-Made Thermostable DNA Polymerase Enables Rapid PCR-Based Detection of Mycoplasma, Ureaplasma and Other Bacteria in the Amniotic Fluid of Preterm Labor Cases.

PubMed

Ueno, Tomohiro; Niimi, Hideki; Yoneda, Noriko; Yoneda, Satoshi; Mori, Masashi; Tabata, Homare; Minami, Hiroshi; Saito, Shigeru; Kitajima, Isao

2015-01-01

Intra-amniotic infection has long been recognized as the leading cause of preterm delivery. Microbial culture is the gold standard for the detection of intra-amniotic infection, but several days are required, and many bacterial species in the amniotic fluid are difficult to cultivate. We developed a novel nested-PCR-based assay for detecting Mycoplasma, Ureaplasma, other bacteria and fungi in amniotic fluid samples within three hours of sample collection. To detect prokaryotes, eukaryote-made thermostable DNA polymerase, which is free from bacterial DNA contamination, is used in combination with bacterial universal primers. In contrast, to detect eukaryotes, conventional bacterially-made thermostable DNA polymerase is used in combination with fungal universal primers. To assess the validity of the PCR assay, we compared the PCR and conventional culture results using 300 amniotic fluid samples. Based on the detection level (positive and negative), 93.3% (280/300) of Mycoplasma, 94.3% (283/300) of Ureaplasma, 89.3% (268/300) of other bacteria and 99.7% (299/300) of fungi matched the culture results. Meanwhile, concerning the detection of bacteria other than Mycoplasma and Ureaplasma, 228 samples were negative according to the PCR method, 98.2% (224/228) of which were also negative based on the culture method. Employing the devised primer sets, mixed amniotic fluid infections of Mycoplasma, Ureaplasma and/or other bacteria could be clearly distinguished. In addition, we also attempted to compare the relative abundance in 28 amniotic fluid samples with mixed infection, and judged dominance by comparing the Ct values of quantitative real-time PCR. We developed a novel PCR assay for the rapid detection of Mycoplasma, Ureaplasma, other bacteria and fungi in amniotic fluid samples. This assay can also be applied to accurately diagnose the absence of bacteria in samples. We believe that this assay will positively contribute to the treatment of intra-amniotic infection and the prevention of preterm delivery.

Eukaryote-Made Thermostable DNA Polymerase Enables Rapid PCR-Based Detection of Mycoplasma, Ureaplasma and Other Bacteria in the Amniotic Fluid of Preterm Labor Cases

PubMed Central

Yoneda, Noriko; Yoneda, Satoshi; Mori, Masashi; Tabata, Homare; Minami, Hiroshi; Saito, Shigeru; Kitajima, Isao

2015-01-01

Background Intra-amniotic infection has long been recognized as the leading cause of preterm delivery. Microbial culture is the gold standard for the detection of intra-amniotic infection, but several days are required, and many bacterial species in the amniotic fluid are difficult to cultivate. Methods We developed a novel nested-PCR-based assay for detecting Mycoplasma, Ureaplasma, other bacteria and fungi in amniotic fluid samples within three hours of sample collection. To detect prokaryotes, eukaryote-made thermostable DNA polymerase, which is free from bacterial DNA contamination, is used in combination with bacterial universal primers. In contrast, to detect eukaryotes, conventional bacterially-made thermostable DNA polymerase is used in combination with fungal universal primers. To assess the validity of the PCR assay, we compared the PCR and conventional culture results using 300 amniotic fluid samples. Results Based on the detection level (positive and negative), 93.3% (280/300) of Mycoplasma, 94.3% (283/300) of Ureaplasma, 89.3% (268/300) of other bacteria and 99.7% (299/300) of fungi matched the culture results. Meanwhile, concerning the detection of bacteria other than Mycoplasma and Ureaplasma, 228 samples were negative according to the PCR method, 98.2% (224/228) of which were also negative based on the culture method. Employing the devised primer sets, mixed amniotic fluid infections of Mycoplasma, Ureaplasma and/or other bacteria could be clearly distinguished. In addition, we also attempted to compare the relative abundance in 28 amniotic fluid samples with mixed infection, and judged dominance by comparing the Ct values of quantitative real-time PCR. Conclusions We developed a novel PCR assay for the rapid detection of Mycoplasma, Ureaplasma, other bacteria and fungi in amniotic fluid samples. This assay can also be applied to accurately diagnose the absence of bacteria in samples. We believe that this assay will positively contribute to the treatment of intra-amniotic infection and the prevention of preterm delivery. PMID:26042418

On the way to identify microorganisms in drinking water distribution networks via DNA analysis of the gut content of freshwater isopods.

PubMed

Mayer, Michael; Keller, Adrian; Szewzyk, Ulrich; Warnecke, Hans-Joachim

2015-05-10

Pure drinking water is the basis for a healthy society. In Germany the drinking water regulations demand for analysis of water via detection of certain microbiological parameters by cultivation only. However, not all prokaryotes can be detected by these standard methods. How to gain more and better information about the bacteria present in drinking water and its distribution systems? The biofilms in drinking water distribution systems are built by bacteria and therefore represent a valuable source of information about the species present. Unfortunately, these biofilms are badly accessible. We thus exploited the circumstance that a lot of metazoans graze the biofilms, so that the content of their guts partly reflects the respective biofilm biocenosis. Therefore, we collected omnivorous isopods, prepared their guts and examined and characterized their contents based on 16S und 18S rDNA analysis. These molecularbiological investigations provide a profound basis for the characterization of the biocenosis and thereby biologically assess the drinking water ecosystems. Combined with a thorough identification of the species and the knowledge of their habitats, this approach can provide useful indications for the assessment of drinking-water quality and the early detection of problems in the distribution system. Copyright © 2015 Elsevier B.V. All rights reserved.

Prevalence of tetracycline resistance genes among multi-drug resistant bacteria from selected water distribution systems in southwestern Nigeria.

PubMed

Adesoji, Ayodele T; Ogunjobi, Adeniyi A; Olatoye, Isaac O; Call, Douglas R; Douglas, Douglas R

2015-06-25

Antibiotic resistance genes [ARGs] in aquatic systems have drawn increasing attention they could be transferred horizontally to pathogenic bacteria. Water treatment plants (WTPs) are intended to provide quality and widely available water to the local populace they serve. However, WTPs in developing countries may not be dependable for clean water and they could serve as points of dissemination for antibiotic resistant bacteria. Only a few studies have investigated the occurrence of ARGs among these bacteria including tetracycline resistance genes in water distribution systems in Nigeria. Multi-drug resistant (MDR) bacteria, including resistance to tetracycline, were isolated from treated and untreated water distribution systems in southwest Nigeria. MDR bacteria were resistant to >3 classes of antibiotics based on break-point assays. Isolates were characterized using partial 16S rDNA sequencing and PCR assays for six tetracycline-resistance genes. Plasmid conjugation was evaluated using E. coli strain DH5α as the recipient strain. Out of the 105 bacteria, 85 (81 %) and 20 (19 %) were Gram- negative or Gram- positive, respectively. Twenty-nine isolates carried at least one of the targeted tetracycline resistance genes including strains of Aeromonas, Alcaligenes, Bacillus, Klebsiella, Leucobacter, Morganella, Proteus and a sequence matching a previously uncultured bacteria. Tet(A) was the most prevalent (16/29) followed by tet(E) (4/29) and tet30 (2/29). Tet(O) was not detected in any of the isolates. Tet(A) was mostly found with Alcaligenes strains (9/10) and a combination of more than one resistance gene was observed only amongst Alcaligenes strains [tet(A) + tet30 (2/10), tet(A) + tet(E) (3/10), tet(E) + tet(M) (1/10), tet(E) + tet30 (1/10)]. Tet(A) was transferred by conjugation for five Alcaligenes and two E. coli isolates. This study found a high prevalence of plasmid-encoded tet(A) among Alcaligenes isolates, raising the possibility that this strain could shuttle resistance plasmids to pathogenic bacteria.

Event extraction of bacteria biotopes: a knowledge-intensive NLP-based approach

PubMed Central

2012-01-01

Background Bacteria biotopes cover a wide range of diverse habitats including animal and plant hosts, natural, medical and industrial environments. The high volume of publications in the microbiology domain provides a rich source of up-to-date information on bacteria biotopes. This information, as found in scientific articles, is expressed in natural language and is rarely available in a structured format, such as a database. This information is of great importance for fundamental research and microbiology applications (e.g., medicine, agronomy, food, bioenergy). The automatic extraction of this information from texts will provide a great benefit to the field. Methods We present a new method for extracting relationships between bacteria and their locations using the Alvis framework. Recognition of bacteria and their locations was achieved using a pattern-based approach and domain lexical resources. For the detection of environment locations, we propose a new approach that combines lexical information and the syntactic-semantic analysis of corpus terms to overcome the incompleteness of lexical resources. Bacteria location relations extend over sentence borders, and we developed domain-specific rules for dealing with bacteria anaphors. Results We participated in the BioNLP 2011 Bacteria Biotope (BB) task with the Alvis system. Official evaluation results show that it achieves the best performance of participating systems. New developments since then have increased the F-score by 4.1 points. Conclusions We have shown that the combination of semantic analysis and domain-adapted resources is both effective and efficient for event information extraction in the bacteria biotope domain. We plan to adapt the method to deal with a larger set of location types and a large-scale scientific article corpus to enable microbiologists to integrate and use the extracted knowledge in combination with experimental data. PMID:22759462

Event extraction of bacteria biotopes: a knowledge-intensive NLP-based approach.

PubMed

Ratkovic, Zorana; Golik, Wiktoria; Warnier, Pierre

2012-06-26

Bacteria biotopes cover a wide range of diverse habitats including animal and plant hosts, natural, medical and industrial environments. The high volume of publications in the microbiology domain provides a rich source of up-to-date information on bacteria biotopes. This information, as found in scientific articles, is expressed in natural language and is rarely available in a structured format, such as a database. This information is of great importance for fundamental research and microbiology applications (e.g., medicine, agronomy, food, bioenergy). The automatic extraction of this information from texts will provide a great benefit to the field. We present a new method for extracting relationships between bacteria and their locations using the Alvis framework. Recognition of bacteria and their locations was achieved using a pattern-based approach and domain lexical resources. For the detection of environment locations, we propose a new approach that combines lexical information and the syntactic-semantic analysis of corpus terms to overcome the incompleteness of lexical resources. Bacteria location relations extend over sentence borders, and we developed domain-specific rules for dealing with bacteria anaphors. We participated in the BioNLP 2011 Bacteria Biotope (BB) task with the Alvis system. Official evaluation results show that it achieves the best performance of participating systems. New developments since then have increased the F-score by 4.1 points. We have shown that the combination of semantic analysis and domain-adapted resources is both effective and efficient for event information extraction in the bacteria biotope domain. We plan to adapt the method to deal with a larger set of location types and a large-scale scientific article corpus to enable microbiologists to integrate and use the extracted knowledge in combination with experimental data.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Larson, Richard; Branch, Darren; Edwards, Thayne

The acoustic wave biosensor is innovative device that is a handheld, battery-powered, portable detection system capable of multiplex identification of a wide range of medically relevant pathogens and their biomolecular signatures — viruses, bacteria, proteins, and DNA — at clinically relevant levels. This detection occurs within minutes — not hours — at the point of care, whether that care is in a physician's office, a hospital bed, or at the scene of a biodefense or biomedical emergency.

An Acoustic Plate Mode Sensor for Biowarfare Toxins, Phase II

DTIC Science & Technology

1997-10-01

Biological agents -- such as bacteria , bacterial toxins and viruses -- must be detected rapidly to allow their neutralization or the quick treatment of...Results were comparable. 16 * r Cyclic voltammetry (CV) studies indicate that the monolayers made with the thiodialkyne, D1, which have been photolyzed...Microprocessor system development is ahead of schedule. Preliminary biosensor data is offered using Y. Pestis. The data indi- cates marginal detection

Microfluidic Transducer for Detecting Nanomechanical Movements of Bacteria

NASA Astrophysics Data System (ADS)

Kara, Vural; Ekinci, Kamil

2017-11-01

Various nanomechanical movements of bacteria are currently being explored as an indication of bacterial viability. Most notably, these movements have been observed to subside rapidly and dramatically when the bacteria are exposed to an effective antibiotic. This suggests that monitoring bacterial movements, if performed with high fidelity, can offer a path to various clinical microbiological applications, including antibiotic susceptibility tests. Here, we introduce a robust and sensitive microfluidic transduction technique for detecting the nanomechanical movements of bacteria. The technique is based on measuring the electrical fluctuations in a microchannel which the bacteria populate. These electrical fluctuations are caused by the swimming of motile, planktonic bacteria and random oscillations of surface-immobilized bacteria. The technique provides enough sensitivity to detect even the slightest movements of a single cell and lends itself to smooth integration with other microfluidic methods and devices; it may eventually be used for rapid antibiotic susceptibility testing. We acknowledge support from Boston University Office of Technology Development, Boston University College of Engineering, NIH (1R03AI126168-01) and The Wallace H. Coulter Foundation.

Quartz crystal microbalance (QCM) as biosensor for the detecting of Escherichia coli O157:H7

NASA Astrophysics Data System (ADS)

Thanh Ngo, Vo Ke; Giang Nguyen, Dang; Phuong Uyen Nguyen, Hoang; Tran, Van Man; Nguyen, Thi Khoa My; Phat Huynh, Trong; Lam, Quang Vinh; Dat Huynh, Thanh; Truong, Thi Ngoc Lien

2014-12-01

Although Escherichia coli (E. coli) is a commensalism organism in the intestine of humans and warm-blooded animals, it can be toxic at higher density and causes diseases, especially the highly toxic E. coli O157:H7. In this paper a quartz crystal microbalance (QCM) biosensor was developed for the detection of E. coli O157:H7 bacteria. The anti-E. coli O157:H7 antibodies were immobilized on a self-assembly monolayer (SAM) modified 5 MHz AT-cut quartz crystal resonator. The SAMs were activated with 16-mercaptopropanoic acid, in the presence of 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC) and ester N-hydroxysuccinimide (NHS). The result of changing frequency due to the adsorption of E. coli O157:H7 was measured by the QCM biosensor system designed and fabricated by ICDREC-VNUHCM. This system gave good results in the range of 102-107 CFU mL-1 E. coli O157:H7. The time of bacteria E. coli O157:H7 detection in the sample was about 50 m. Besides, QCM biosensor from SAM method was comparable to protein A method-based piezoelectric immunosensor in terms of the amount of immobilized antibodies and detection sensitivity.

Common Virulence Factors and Tissue Targets of Entomopathogenic Bacteria for Biological Control of Lepidopteran Pests

PubMed Central

Castagnola, Anaïs; Stock, S. Patricia

2014-01-01

This review focuses on common insecticidal virulence factors from entomopathogenic bacteria with special emphasis on two insect pathogenic bacteria Photorhabdus (Proteobacteria: Enterobacteriaceae) and Bacillus (Firmicutes: Bacillaceae). Insect pathogenic bacteria of diverse taxonomic groups and phylogenetic origin have been shown to have striking similarities in the virulence factors they produce. It has been suggested that the detection of phage elements surrounding toxin genes, horizontal and lateral gene transfer events, and plasmid shuffling occurrences may be some of the reasons that virulence factor genes have so many analogs throughout the bacterial kingdom. Comparison of virulence factors of Photorhabdus, and Bacillus, two bacteria with dissimilar life styles opens the possibility of re-examining newly discovered toxins for novel tissue targets. For example, nematodes residing in the hemolymph may release bacteria with virulence factors targeting neurons or neuromuscular junctions. The first section of this review focuses on toxins and their context in agriculture. The second describes the mode of action of toxins from common entomopathogens and the third draws comparisons between Gram positive and Gram negative bacteria. The fourth section reviews the implications of the nervous system in biocontrol. PMID:24634779

A fully automated microfluidic-based electrochemical sensor for real-time bacteria detection.

PubMed

Altintas, Zeynep; Akgun, Mete; Kokturk, Guzin; Uludag, Yildiz

2018-02-15

A fully automated microfluidic-based electrochemical biosensor was designed and manufactured for pathogen detection. The quantification of Escherichia coli was investigated with standard and nanomaterial amplified immunoassays in the concentration ranges of 0.99 × 10 4 3.98 × 10 9 cfu mL -1 and 103.97 × 10 7 cfu mL -1 which resulted in detection limits of 1.99 × 10 4 cfu mL -1 and 50 cfu mL -1 , respectively. The developed methodology was then applied for E. coli quantification in water samples using nanomaterial modified assay. Same detection limit for E. coli was achieved for real sample analysis with a little decrease on the sensor signal. Cross-reactivity studies were conducted by testing Shigella, Salmonella spp., Salmonella typhimurium and Staphylococcus aureus on E. coli specific antibody surface that confirmed the high specificity of the developed immunoassays. The sensor surface could be regenerated multiple times which significantly reduces the cost of the system. Our custom-designed biosensor is capable of detecting bacteria with high sensitivity and specificity, and can serve as a promising tool for pathogen detection. Copyright © 2017 Elsevier B.V. All rights reserved.

Microfluidics microFACS for Life Detection

NASA Technical Reports Server (NTRS)

Platt, Donald W.; Hoover, Richard B.

2010-01-01

A prototype micro-scale Fluorescent Activated Cell Sorter (microFACS) for life detection has been built and is undergoing testing. A functional miniature microfluidics instrument with the ability to remotely distinguish live or dead bacterial cells from abiotic particulates in ice or permafrost of icy bodies of the solar system would be of fundamental value to NASA. The use of molecular probes to obtain the bio-signature of living or dead cells could answer the most fundamental question of Astrobiology: Does life exist beyond Earth? The live-dead fluorescent stains to be used in the microFACS instrument function only with biological cell walls. The detection of the cell membranes of living or dead bacteria (unlike PAH's and many other Biomarkers) would provide convincing evidence of present or past life. This miniature device rapidly examine large numbers of particulates from a polar ice or permafrost sample and distinguish living from dead bacteria cells and biological cells from mineral grains and abiotic particulates and sort the cells and particulates based on a staining system. Any sample found to exhibit fluorescence consistent with living cells could then be used in conjunction with a chiral labeled release experiment or video microscopy system to seek addition evidence for cellular metabolism or motility. Results of preliminary testing and calibration of the microFACS prototype instrument system with pure cultures and enrichment assemblages of microbial extremophiles will be reported.

Occurrence of low molecular weight thiols in biological systems

NASA Technical Reports Server (NTRS)

Fahey, Robert C.; Newton, Gerald L.

1983-01-01

Bromobimane labeling and high performance chromatography analysis were applied to various species of bacteria, plant tissues, and animal tissues. The reaction between thiols and monobromobimane is studied. Chromatograms revealing peaks produced by nonthiols and thiols are analyzed and compared. It is observed that all the bacteria species contain hydrogen sulfide, and glutathione is contained in facultative and aerobic gram-negative bacteria. For the plant tissues, the data reveal that mung bean sprouts contain homoglutathione and no glutathione; alfalfa sprouts contain homoglutathione and glutathione; the pea seed, nonlegumes, and fungi contain glutathione and no homoglutathione. It is detected that the main thiol in the animal tissues is glutathione. Based on the data, it is suggested that glutathione has an essential function in higher organisms.

Method of Detecting Coliform Bacteria from Reflected Light

NASA Technical Reports Server (NTRS)

Vincent, Robert K. (Inventor)

2014-01-01

The present invention relates to a method of detecting coliform bacteria in water from reflected light, and also includes devices for the measurement, calculation and transmission of data relating to that method.

Adhesion of Mycobacterium smegmatis to Charged Surfaces and Diagnostics Implications

NASA Astrophysics Data System (ADS)

Gorse, Diane; Dhinojwala, Ali; Moore, Francisco

Pulmonary tuberculosis (PTB) causes more than 1 million deaths annually. Smear microscopy is a primary rapid detection tool in areas where 95 % of PTB cases occur. This technique, in which the sputum of a symptomatic patient is stained and examined using a light microscope for Mycobacterium tuberculosis (MTB) shows sensitivity between 20 and 60 %. Insufficient bacterial isolation during sample preparation may be a reason for low sensitivity. We are optimizing a system to capture bacteria on the basis of electrostatic interactions to more thoroughly isolate bacteria from suspension and facilitate more accurate detection. Silica supports coated with positively-charged polyelectrolyte, poly(diallyldimethylammonium chloride), captured approximately 4.1 times more Mycobacterium smegmatis, a model organism for MTB, than was captured on negatively-charged silica substrates. Future experimentation will employ branched polymer systems and seek to justify the use of colloidal stability theories to describe initial capture. Supported by University of Akron, Department of Polymer Science, Department of Biology; LORD Corporation.

Characterizing the Purple Earth: Modeling the globally integrated spectral variability of the Archean Earth

DOE Office of Scientific and Technical Information (OSTI.GOV)

Sanromá, E.; Pallé, E.; López, R.

2014-01-01

Ongoing searches for exoplanetary systems have revealed a wealth of planets with diverse physical properties. Planets even smaller than the Earth have already been detected and the efforts of future missions are aimed at the discovery, and perhaps characterization, of small rocky exoplanets within the habitable zone of their stars. Clearly, what we know about our planet will be our guideline for the characterization of such planets. However, the Earth has been inhabited for at least 3.8 Gyr and its appearance has changed with time. Here, we have studied the Earth during the Archean eon, 3.0 Gyr ago. At thatmore » time, one of the more widespread life forms on the planet was purple bacteria. These bacteria are photosynthetic microorganisms and can inhabit both aquatic and terrestrial environments. Here, we use a radiative transfer model to simulate the visible and near-infrared radiation reflected by our planet, taking into account several scenarios regarding the possible distribution of purple bacteria over continents and oceans. We find that purple bacteria have a reflectance spectrum that has a strong reflectivity increase, similar to the red edge of leafy plants, although shifted redward. This feature produces a detectable signal in the disk-averaged spectra of our planet, depending on cloud amount and purple bacteria concentration/distribution. We conclude that by using multi-color photometric observations, it is possible to distinguish between an Archean Earth in which purple bacteria inhabit vast extensions of the planet and a present-day Earth with continents covered by deserts, vegetation, or microbial mats.« less

Automated detection of bacteria in urine

NASA Technical Reports Server (NTRS)

Fleig, A. J.; Picciolo, G. L.; Chappelle, E. W.; Kelbaugh, B. N.

1972-01-01

A method for detecting the presence of bacteria in urine was developed which utilizes the bioluminescent reaction of adenosine triphosphate with luciferin and luciferase derived from the tails of fireflies. The method was derived from work on extraterrestrial life detection. A device was developed which completely automates the assay process.

Quick and simple estimation of bacteria using a fluorescent paracetamol dimer-Au nanoparticle composite

NASA Astrophysics Data System (ADS)

Sahoo, Amaresh Kumar; Sharma, Shilpa; Chattopadhyay, Arun; Ghosh, Siddhartha Sankar

2012-02-01

Rapid, simple and sensitive detection of bacterial contamination is critical for safeguarding public health and the environment. Herein, we report an easy method of detection as well as enumeration of the bacterial cell number on the basis of fluorescence quenching of a non-antibacterial fluorescent nanocomposite, consisting of paracetamol dimer (PD) and Au nanoparticles (NPs), in the presence of bacteria. The composite was synthesized by reaction of paracetamol (p-hydroxyacetanilide) with HAuCl4. The Au NPs of the composite were characterized using UV-Vis spectroscopy, transmission electron microscopy (TEM), X-ray diffraction and selected area electron diffraction analysis. The paracetamol dimer in the composite showed emission peak at 435 nm when excited at 320 nm. The method successfully detected six bacterial strains with a sensitivity of 100 CFU mL-1. The Gram-positive and Gram-negative bacteria quenched the fluorescence of the composite differently, making it possible to distinguish between the two. The TEM analysis showed interaction of the composite with bacteria without any apparent damage to the bacteria. The chi-square test established the accuracy of the method. Quick, non-specific and highly sensitive detection of bacteria over a broad range of logarithmic dilutions within a short span of time demonstrates the potential of this method as an alternative to conventional methods.Rapid, simple and sensitive detection of bacterial contamination is critical for safeguarding public health and the environment. Herein, we report an easy method of detection as well as enumeration of the bacterial cell number on the basis of fluorescence quenching of a non-antibacterial fluorescent nanocomposite, consisting of paracetamol dimer (PD) and Au nanoparticles (NPs), in the presence of bacteria. The composite was synthesized by reaction of paracetamol (p-hydroxyacetanilide) with HAuCl4. The Au NPs of the composite were characterized using UV-Vis spectroscopy, transmission electron microscopy (TEM), X-ray diffraction and selected area electron diffraction analysis. The paracetamol dimer in the composite showed emission peak at 435 nm when excited at 320 nm. The method successfully detected six bacterial strains with a sensitivity of 100 CFU mL-1. The Gram-positive and Gram-negative bacteria quenched the fluorescence of the composite differently, making it possible to distinguish between the two. The TEM analysis showed interaction of the composite with bacteria without any apparent damage to the bacteria. The chi-square test established the accuracy of the method. Quick, non-specific and highly sensitive detection of bacteria over a broad range of logarithmic dilutions within a short span of time demonstrates the potential of this method as an alternative to conventional methods. Electronic supplementary information (ESI) available. See DOI: 10.1039/c2nr11837h

Dual Recognition Strategy for Specific and Sensitive Detection of Bacteria Using Aptamer-Coated Magnetic Beads and Antibiotic-Capped Gold Nanoclusters.

PubMed

Cheng, Dan; Yu, Mengqun; Fu, Fei; Han, Weiye; Li, Gan; Xie, Jianping; Song, Yang; Swihart, Mark T; Song, Erqun

2016-01-05

Food poisoning and infectious diseases caused by pathogenic bacteria such as Staphylococcus aureus (SA) are serious public health concerns. A method of specific, sensitive, and rapid detection of such bacteria is essential and important. This study presents a strategy that combines aptamer and antibiotic-based dual recognition units with magnetic enrichment and fluorescent detection to achieve specific and sensitive quantification of SA in authentic specimens and in the presence of much higher concentrations of other bacteria. Aptamer-coated magnetic beads (Apt-MB) were employed for specific capture of SA. Vancomycin-stabilized fluorescent gold nanoclusters (AuNCs@Van) were prepared by a simple one-step process and used for sensitive quantification of SA in the range of 32-10(8) cfu/mL with the detection limit of 16 cfu/mL via a fluorescence intensity measurement. And using this strategy, about 70 cfu/mL of SA in complex samples (containing 3 × 10(8) cfu/mL of other different contaminated bacteria) could be successfully detected. In comparison to prior studies, the developed strategy here not only simplifies the preparation procedure of the fluorescent probes (AuNCs@Van) to a great extent but also could sensitively quantify SA in the presence of much higher concentrations of other bacteria directly with good accuracy. Moreover, the aptamer and antibiotic used in this strategy are much less expensive and widely available compared to common-used antibodies, making it cost-effective. This general aptamer- and antibiotic-based dual recognition strategy, combined with magnetic enrichment and fluorescent detection of trace bacteria, shows great potential application in monitoring bacterial food contamination and infectious diseases.

Towards optical fibre based Raman spectroscopy for the detection of surgical site infection

NASA Astrophysics Data System (ADS)

Thompson, Alex J.; Koziej, Lukasz; Williams, Huw D.; Elson, Daniel S.; Yang, Guang-Zhong

2016-03-01

Surgical site infections (SSIs) are common post-surgical complications that remain significant clinical problems, as they are associated with substantial mortality and morbidity. As such, there is significant interest in the development of minimally invasive techniques that permit early detection of SSIs. To this end, we are applying a compact, clinically deployable Raman spectrometer coupled to an optical fibre probe to the study of bacteria, with the long term goal of using Raman spectroscopy to detect infection in vivo. Our system comprises a 785 nm laser diode for excitation and a commercial (Ocean Optics, Inc.) Raman spectrometer for detection. Here we discuss the design, optimisation and validation of this system, and describe our first experiences interrogating bacterial cells (Escherichia coli) in vitro.

Scanning electron microscopy coupled with energy-dispersive X-ray spectrometry for quick detection of sulfur-oxidizing bacteria in environmental water samples

NASA Astrophysics Data System (ADS)

Sun, Chengjun; Jiang, Fenghua; Gao, Wei; Li, Xiaoyun; Yu, Yanzhen; Yin, Xiaofei; Wang, Yong; Ding, Haibing

2017-01-01

Detection of sulfur-oxidizing bacteria has largely been dependent on targeted gene sequencing technology or traditional cell cultivation, which usually takes from days to months to carry out. This clearly does not meet the requirements of analysis for time-sensitive samples and/or complicated environmental samples. Since energy-dispersive X-ray spectrometry (EDS) can be used to simultaneously detect multiple elements in a sample, including sulfur, with minimal sample treatment, this technology was applied to detect sulfur-oxidizing bacteria using their high sulfur content within the cell. This article describes the application of scanning electron microscopy imaging coupled with EDS mapping for quick detection of sulfur oxidizers in contaminated environmental water samples, with minimal sample handling. Scanning electron microscopy imaging revealed the existence of dense granules within the bacterial cells, while EDS identified large amounts of sulfur within them. EDS mapping localized the sulfur to these granules. Subsequent 16S rRNA gene sequencing showed that the bacteria detected in our samples belonged to the genus Chromatium, which are sulfur oxidizers. Thus, EDS mapping made it possible to identify sulfur oxidizers in environmental samples based on localized sulfur within their cells, within a short time (within 24 h of sampling). This technique has wide ranging applications for detection of sulfur bacteria in environmental water samples.

Evaluation of the NanoCHIP® Gastrointestinal Panel (GIP) Test for Simultaneous Detection of Parasitic and Bacterial Enteric Pathogens in Fecal Specimens

PubMed Central

Ken Dror, Shifra; Pavlotzky, Elsa; Barak, Mira

2016-01-01

Infectious gastroenteritis is a global health problem associated with high morbidity and mortality rates. Rapid and accurate diagnosis is crucial to allow appropriate and timely treatment. Current laboratory stool testing has a long turnaround time (TAT) and demands highly qualified personnel and multiple techniques. The need for high throughput and the number of possible enteric pathogens compels the implementation of a molecular approach which uses multiplex technology, without compromising performance requirements. In this work we evaluated the feasibility of the NanoCHIP® Gastrointestinal Panel (GIP) (Savyon Diagnostics, Ashdod, IL), a molecular microarray-based screening test, to be used in the routine workflow of our laboratory, a big outpatient microbiology laboratory. The NanoCHIP® GIP test provides simultaneous detection of nine major enteric bacteria and parasites: Campylobacter spp., Salmonella spp., Shigella spp., Giardia sp., Cryptosporidium spp., Entamoeba histolytica, Entamoeba dispar, Dientamoeba fragilis, and Blastocystis spp. The required high-throughput was obtained by the NanoCHIP® detection system together with the MagNA Pure 96 DNA purification system (Roche Diagnostics Ltd., Switzerland). This combined system has demonstrated a higher sensitivity and detection yield compared to the conventional methods in both, retrospective and prospective samples. The identification of multiple parasites and bacteria in a single test also enabled increased efficiency of detecting mixed infections, as well as reduced hands-on time and work load. In conclusion, the combination of these two automated systems is a proper response to the laboratory needs in terms of improving laboratory workflow, turn-around-time, minimizing human errors and can be efficiently integrated in the routine work of the laboratory. PMID:27447173

Sanitary evaluation of domestic water supply facilities with storage tanks and detection of Aeromonas, enteric and related bacteria in domestic water facilities in Okinawa Prefecture of Japan.

PubMed

Miyagi, Kazufumi; Sano, Kouichi; Hirai, Itaru

2017-08-01

To provide for temporary restrictions of the public water supply system, storage tanks are commonly installed in the domestic water systems of houses and apartment buildings in Okinawa Prefecture of Japan. To learn more about the sanitary condition and management of these water supply facilities with storage tanks (hereafter called "storage tank water systems") and the extent of bacterial contamination of water from these facilities, we investigated their usage and the existence of Aeromonas, enteric and related bacteria. Verbal interviews concerning the use and management of the storage tank water systems were carried out in each randomly sampled household. A total of 54 water samples were collected for bacteriological and physicochemical examinations. Conventional methods were used for total viable count, fecal coliforms, identification of bacteria such as Aeromonas, Enterobacteriaceae and non-fermentative Gram-negative rods (NF-GNR), and measurement of residual chlorine. On Aeromonas species, tests for putative virulence factor and an identification using 16S rRNA and rpoB genes were also performed. Water from the water storage systems was reported to be consumed directly without boiling in 22 of the 54 houses (40.7%). 31 of the sampled houses had installed water storage tanks of more than 1 cubic meter (m 3 ) per inhabitant, and in 21 of the sampled houses, the tank had never been cleaned. In all samples, the total viable count and fecal coliforms did not exceed quality levels prescribed by Japanese waterworks law. Although the quantity of bacteria detected was not high, 23 NF-GNR, 14 Enterobacteriaceae and 5 Aeromonas were isolated in 42.6%, 7.4% and 3.7% of samples respectively. One isolated A. hydrophila and four A. caviae possessed various putative virulence factors, especially A. hydrophila which had diverse putative pathogenic genes such as aer, hlyA, act, alt, ast, ser, and dam. Many bacteria were isolated when the concentration of residual chlorine was below 0.1 mg/l and the water temperature was above 20 °C. These results suggest that elevated water temperature and mismatch between tank size and water demand lead to loss of residual chlorine in tap water. Therefore, to minimize growth of aquatic bacteria such as Aeromonas spp. and Pseudomonas spp., we recommend that an appropriate size tank and/or volume of stored water is always used, and also suggest installation of some means of reducing water temperature such as shading. Copyright © 2017 Elsevier Ltd. All rights reserved.

Detection and inhibition of bacterial cell-cell communication.

PubMed

Rice, Scott A; McDougald, Diane; Givskov, Michael; Kjelleberg, Staffan

2008-01-01

Bacteria communicate with other members of their community through the secretion and perception of small chemical cues or signals. The recognition of a signal normally leads to the expression of a large suite of genes, which in some bacteria are involved in the regulation of virulence factors, and as a result, these signaling compounds are key regulatory factors in many disease processes. Thus, it is of interest when studying pathogens to understand the mechanisms used to control the expression of virulence genes so that strategies might be devised for the control of those pathogens. Clearly, the ability to interfere with this process of signaling represents a novel approach for the treatment of bacterial infections. There is a broad range of compounds that bacteria can use for signaling purposes, including fatty acids, peptides, N-acylated homoserine lactones, and the signals collectively called autoinducer 2 (AI-2). This chapter will focus on the latter two signaling systems as they are present in a range of medically relevant bacteria, and here we describe assays for determining whether an organism produces a particular signal and assays that can be used to identify inhibitors of the signaling cascade. Lastly, the signal detection and inhibition assays will be directly linked to the expression of virulence factors of specific pathogens.

Simultaneous bioremediation and biodetection of mercury ion through surface display of carboxylesterase E2 from Pseudomonas aeruginosa PA1.

PubMed

Yin, Kun; Lv, Min; Wang, Qiaoning; Wu, Yixuan; Liao, Chunyang; Zhang, Weiwei; Chen, Lingxin

2016-10-15

Mercury is a toxic heavy metal and presents significant threats to organisms and natural ecosystems. Recently, the mercury remediation as well as its detection by environmental-friendly biotechnology has received increasing attention. In this study, carboxylesterase E2 from mercury-resistant strain Pseudomonas aeruginosa PA1 has been successfully displayed on the outer membrane of Escherichia coli Top10 bacteria to simultaneously adsorb and detect mercury ion (Hg(2+)). The transmission electron microscopy analysis shows that Hg(2+) can be absorbed by carboxylesterase E2 and accumulated on the outer membrane of surface-displayed E. coli bacteria. The adsorption of Hg(2+) followed a physicochemical, equilibrated and saturatable mechanism, which well fits the traditional Langmuir adsorption model. The surface-displayed system can be regenerated through regulating pH values. As its activity can be inhibited by Hg(2+), carboxylesterase E2 has been used to detect the concentration of Hg(2+) in water samples. The developed surface display system will be of great potential in the simultaneous bioremediation and biodetection of environmental mercury pollution. Copyright © 2016 Elsevier Ltd. All rights reserved.

Detection of Antibiotics and Evaluation of Antibacterial Activity with Screen-Printed Electrodes

PubMed Central

Titoiu, Ana Maria; Marty, Jean-Louis

2018-01-01

This review provides a brief overview of the fabrication and properties of screen-printed electrodes and details the different opportunities to apply them for the detection of antibiotics, detection of bacteria and antibiotic susceptibility. Among the alternative approaches to costly chromatographic or ELISA methods for antibiotics detection and to lengthy culture methods for bacteria detection, electrochemical biosensors based on screen-printed electrodes present some distinctive advantages. Chemical and (bio)sensors for the detection of antibiotics and assays coupling detection with screen-printed electrodes with immunomagnetic separation are described. With regards to detection of bacteria, the emphasis is placed on applications targeting viable bacterial cells. While the electrochemical sensors and biosensors face many challenges before replacing standard analysis methods, the potential of screen-printed electrodes is increasingly exploited and more applications are anticipated to advance towards commercial analytical tools. PMID:29562637

Microfluidics: innovative approaches for rapid diagnosis of antibiotic-resistant bacteria.

PubMed

Aroonnual, Amornrat; Janvilisri, Tavan; Ounjai, Puey; Chankhamhaengdecha, Surang

2017-02-28

The emergence of antibiotic-resistant bacteria has become a major global health concern. Rapid and accurate diagnostic strategies to determine the antibiotic susceptibility profile prior to antibiotic prescription and treatment are critical to control drug resistance. The standard diagnostic procedures for the detection of antibiotic-resistant bacteria, which rely mostly on phenotypic characterization, are time consuming, insensitive and often require skilled personnel, making them unsuitable for point-of-care (POC) diagnosis. Various molecular techniques have therefore been implemented to help speed up the process and increase sensitivity. Over the past decade, microfluidic technology has gained great momentum in medical diagnosis as a series of fluid handling steps in a laboratory can be simplified and miniaturized on to a small platform, allowing marked reduction of sample amount, high portability and tremendous possibility for integration with other detection technologies. These advantages render the microfluidic system a great candidate to be developed into an easy-to-use sample-to-answer POC diagnosis suitable for application in remote clinical settings. This review provides an overview of the current development of microfluidic technologies for the nucleic acid based and phenotypic-based detections of antibiotic resistance. © 2017 The Author(s). Published by Portland Press Limited on behalf of the Biochemical Society.

Detection of Alicyclobacillus species in fruit juice using a random genomic DNA microarray chip.

PubMed

Jang, Jun Hyeong; Kim, Sun-Joong; Yoon, Bo Hyun; Ryu, Jee-Hoon; Gu, Man Bock; Chang, Hyo-Ihl

2011-06-01

This study describes a method using a DNA microarray chip to rapidly and simultaneously detect Alicyclobacillus species in orange juice based on the hybridization of genomic DNA with random probes. Three food spoilage bacteria were used in this study: Alicyclobacillus acidocaldarius, Alicyclobacillus acidoterrestris, and Alicyclobacillus cycloheptanicus. The three Alicyclobacillus species were adjusted to 2 × 10(3) CFU/ml and inoculated into pasteurized 100% pure orange juice. Cy5-dCTP labeling was used for reference signals, and Cy3-dCTP was labeled for target genomic DNA. The molar ratio of 1:1 of Cy3-dCTP and Cy5-dCTP was used. DNA microarray chips were fabricated using randomly fragmented DNA of Alicyclobacillus spp. and were hybridized with genomic DNA extracted from Bacillus spp. Genomic DNA extracted from Alicyclobacillus spp. showed a significantly higher hybridization rate compared with DNA of Bacillus spp., thereby distinguishing Alicyclobacillus spp. from Bacillus spp. The results showed that the microarray DNA chip containing randomly fragmented genomic DNA was specific and clearly identified specific food spoilage bacteria. This microarray system is a good tool for rapid and specific detection of thermophilic spoilage bacteria, mainly Alicyclobacillus spp., and is useful and applicable to the fruit juice industry.

Detection of heavy metal resistance bioluminescence bacteria using microplate bioassay method.

PubMed

Ranjitha, P; Karthy, E S

2012-01-01

Effects of different heavy metals on Vibrio harveyi, V. fischeri, Photobacterium phosphoreum and P. leiognathi were examined. Checkerboard assay was used for the detection of the natural metal tolerance levels of a large number of marine luminous eubacteria. 57 strains of luminous bacteria were investigated for their natural patterns of heavy metal tolerance. The behaviors of these strains were not homogeneous with respect to all metals tested, even within the strains belonging to the same genus. At least 1 to 4 different MICs were detected for every metal except barium and cobalt. Isolated bacteria were tested for the presence of plasmids using the modified alkaline lysis method, was effective for identification of plasmids of different sizes. This study revealed the frequency of the occurrence of plasmids in heavy metal resistance bacteria and inferred that plasmids are highly ubiquitous and predominant in most heavy metal resistant bacteria.

Bacterial community of iron tubercles from a drinking water distribution system and its occurrence in stagnant tap water.

PubMed

Chen, Lu; Jia, Rui-Bao; Li, Li

2013-07-01

Bacteria in drinking water distribution systems can cause deterioration of the water quality, and the microbial quality of tap water is closely related to consumer health. In the present study, the potential effects of bacteria attached to cast iron pipes on tap water in a distribution system were investigated. Comparison of the bacterial community composition of pipe tubercles with that of stagnant tap water samples based on a denaturing gradient gel electrophoresis analysis of the 16S rRNA gene revealed that the communities were related. Specifically, the main bacterial members were identical to each other. The bacterial community was found to be dominated by Firmicutes, Actinobacteria, and Proteobacteria, which included Rhizobium, Pseudomonas, Lactococcus, Brevundimonas, Rheinheimera, Arthrobacter, Bacillus, and Herbaspirillum. Heterotrophic bacteria proliferation was observed during the period of stagnation, followed by a decrease of assimilable organic carbon and a slight increase of microbially available phosphorus. These findings indicated that the regrowth of bacteria might be boosted by the release of nutrients such as phosphorus from the pipe walls, as well as the decline of residual chlorine during stagnation. Inorganic contaminants at low levels, including Al, Mn, Zn, Pb, Cr, Cu, and Ni, were detected in tubercles and were concentrated in particulates from tap water following the release of iron during stagnation.

Engineering Bacterial Thiosulfate and Tetrathionate Sensors for Detecting Gut Inflammation

DTIC Science & Technology

2017-04-03

Article Engineering bacterial thiosulfate and tetrathionate sensors for detecting gut inflammation Kristina N-M Daeffler1 , Jeffrey D Galley2, Ravi U...interest in using genetically engineered sensor bacteria to study gut microbiota pathways, and diagnose or treat associated diseases. Here, we...understood. Genetically engineered sensor bacteria have untapped potential as tools for analyzing gut pathways. Bacteria have evolved sensors of a large

Development of a multiplex PCR assay for rapid and simultaneous detection of four genera of fish pathogenic bacteria.

PubMed

Zhang, D F; Zhang, Q Q; Li, A H

2014-11-01

Species of genus Aeromonas, Vibrio, Edwardsiella and Streptococcus are the most common fish pathogenic bacteria that cause economically devastating losses in aquaculture. A multiplex polymerase chain reaction (mPCR) was developed for the simultaneous detection and differentiation of the four genera of fish pathogenic bacteria. Through the use of genus-specific primers instead of species-specific ones, the current mPCR covered much more target bacterial species compared with previously reported species-specific mPCR methods. The specificity of the four putative genus-specific primers was validated experimentally while used exclusively (uniplex PCR) or combined (mPCR) against bacterial genomic DNA templates of the target bacteria and nontarget bacteria. The PCR amplicons for the following genera were obtained as expected: Aeromonas (875 bp), Vibrio (524 bp), Edwardsiella (302 bp) and Streptococcus (197 bp), and the fragments could be separated clearly on the agarose gel electrophoresis. The mPCR did not produce nonspecific amplification products when used to amplify 21 nontarget species of bacteria. The mPCR detection limits for each target bacterial genera were 50 colony-forming units (CFU) in pure culture and 100 CFU in fish tissue samples. In conclusion, the mPCR assay was proven to be a powerful alternative to the conventional culture-based method, given its rapid, specific, sensitive and reliable detection of target pathogens. The fish pathogenic bacteria of genus Aeromonas, Vibrio, Edwardsiella and Streptococcus frequently cause severe outbreaks of diseases in cultured fish, and the genus-specific multiplex PCR assay developed in this study can detect the bacteria of the four genera when present in the samples either alone or mixed. The mPCR assay is expected to identify the causative agents more efficiently than uniplex PCR or species-specific multiplex PCR for clinical diagnosis, resulting in the earlier implementation of control measures. This mPCR assay provides a rapid, specific and sensitive tool for the detection or identification of common fish pathogenic bacteria in aquaculture practice. © 2014 The Society for Applied Microbiology.

A novel multifunctional electrochemical platform for simultaneous detection, elimination, and inactivation of pathogenic bacteria based on the Vancomycin-functionalised AgNPs/3D-ZnO nanorod arrays.

PubMed

Yang, Zhiqing; Wang, Yi; Zhang, Dun

2017-12-15

A novel fast, sensitive, and specific multifunctional electrochemical platform has been proposed for simultaneous detection, elimination, and inactivation of pathogenic bacteria for the first time. The platform is constituted with three-dimensional ZnO nanorod arrays (3D-ZnO) decorated with sliver nanoparticles (AgNPs) and functionalized with vancomycin (Van). Based on the specific recognition of Van for Gram-positive bacteria, the fabricated electrochemical platform has presented high detection sensitivity to Staphylococcus aureus with a low detection limit of 330cfu/mL and adaptable bacterial-elimination efficiency (50%) at low concentrations (1000-2000cfu/mL). Moreover, the platform has shown high antibacterial activity (99.99%) arising from the synergistic germicidal effect of the composited antibacterial AgNPs and Van units. The current work could provide new strategies to construct advanced platforms for simultaneous detection, elimination, and inactivation of various pathogenic bacteria. Copyright © 2017 Elsevier B.V. All rights reserved.

Bacterial detection using bacteriophages and gold nanorods by following time-dependent changes in Raman spectral signals.

PubMed

Moghtader, Farzaneh; Tomak, Aysel; Zareie, Hadi M; Piskin, Erhan

2018-03-27

This study attemps to develop bacterial detection strategies using bacteriophages and gold nanorods (GNRs) by Raman spectral analysis. Escherichia coli was selected as the target and its specific phage was used as the bioprobe. Target bacteria and phages were propagated/purified by traditional techniques. GNRs were synthesized by using hexadecyltrimethyl ammonium bromide (CTAB) as stabilizer. A two-step detection strategy was applied: Firstly, the target bacteria were interacted with GNRs in suspensions, and then they were dropped onto silica substrates for detection. It was possible to obtain clear surface-enchanced Raman spectroscopy (SERS) peaks of the target bacteria, even without using phages. In the second step, the phage nanoemulsions were droped onto the bacterial-GNRs complexes on those surfaces and time-dependent changes in the Raman spectra were monitored at different time intervals upto 40 min. These results demonstrated that how one can apply phages with plasmonic nanoparticles for detection of pathogenic bacteria very effectively in a quite simple test.

Characterization of carotenoids in soil bacteria and investigation of their photodegradation by UVA radiation via resonance Raman spectroscopy.

PubMed

Kumar B N, Vinay; Kampe, Bernd; Rösch, Petra; Popp, Jürgen

2015-07-07

A soil habitat consists of an enormous number of pigmented bacteria with the pigments mainly composed of diverse carotenoids. Most of the pigmented bacteria in the top layer of the soil are photoprotected from exposure to huge amounts of UVA radiation on a daily basis by these carotenoids. The photostability of these carotenoids depends heavily on the presence of specific features like a carbonyl group or an ionone ring system on its overall structure. Resonance Raman spectroscopy is one of the most sensitive and powerful techniques to detect and characterize these carotenoids and also monitor processes associated with them in their native system at a single cell resolution. However, most of the resonance Raman profiles of carotenoids have very minute differences, thereby making it extremely difficult to confirm if these differences are attributed to the presence of different carotenoids or if it is a consequence of their interaction with other cellular components. In this study, we devised a method to overcome this problem by monitoring also the photodegradation of the carotenoids in question by UVA radiation wherein a differential photodegradation response will confirm the presence of different carotenoids irrespective of the proximities in their resonance Raman profiles. Using this method, the detection and characterization of carotenoids in pure cultures of five species of pigmented coccoid soil bacteria is achieved. We also shed light on the influence of the structure of the carotenoid on its photodegradation which can be exploited for use in the characterization of carotenoids via resonance Raman spectroscopy.

Aptamer-based hydrogel barcodes for the capture and detection of multiple types of pathogenic bacteria.

PubMed

Xu, Yueshuang; Wang, Huan; Luan, Chengxin; Liu, Yuxiao; Chen, Baoan; Zhao, Yuanjin

2018-02-15

Rapid and sensitive diagnosing hematological infections based on the separation and detection of pathogenic bacteria in the patient's blood is a significant challenge. To address this, we herein present a new barcodes technology that can simultaneously capture and detect multiple types of pathogenic bacteria from a complex sample. The barcodes are poly (ethylene glycol) (PEG) hydrogel inverse opal particles with characteristic reflection peak codes that remain stable during bacteria capture on their surfaces. As the spherical surface of the particles has ordered porous nanostructure, the barcodes can provide not only more surface area for probe immobilization and reaction, but also a nanopatterned platform for highly efficient bioreactions. In addition, the PEG hydrogel scaffold could decrease the non-specificity adsorption by its anti-adhesive effect, and the decorated aptamer probes in the scaffolds could increase the sensitivity, reliability, and specificity of the bacteria capture and detection. Moreover, the tagged magnetic nanoparticles in the PEG scaffold could impart the barcodes with controllable movement under magnetic fields, which can be used to significantly increase the reaction speed and simplify the processing of the bioassays. Based on the describe barcodes, it was demonstrated that the bacteria could be captured and identified even at low bacterial concentrations (100 CFU mL -1 ) within 2.5h, which is effectively shortened in comparison with the "gold standard" in clinic. These features make the barcodes ideal for capturing and detecting multiple bacteria from clinical samples for hematological infection diagnostics. Copyright © 2017 Elsevier B.V. All rights reserved.

Keeping the Air Clean and Safe: An Anthrax Smoke Detector

NASA Technical Reports Server (NTRS)

2005-01-01

Scientists at work in the Planetary Protection division at NASA s Jet Propulsion Laboratory (JPL) sterilize everything before blasting it to the Red Planet. They take great pains to ensure that all spacecraft are void of bacterial life, especially the microscopic bacteria that can live hundreds of years in their spore states. No one is quite sure what Earthly germs would do on Mars, but scientists agree that it is safest to keep the Martian terrain as undisturbed as possible. Errant Earth germs would also render useless the instruments placed on exploration rovers to look for signs of life, as the life that they registered would be life that came with them from Earth. A team at JPL, headed by Dr. Adrian Ponce, developed a bacterial spore-detection system that uses a simple and robust chemical reaction that visually alerts Planetary Protection crews. It is a simple air filter that traps micron-sized bacterial spores and then submits them to the chemical reaction. When the solution is then viewed under an ultraviolet light, the mixture will glow green if it is contaminated by bacteria. Scientists can then return to the scrubbing and cleaning stages of the sterilization process to remove these harmful bacteria. The detection system is the space-bound equivalent of having your hands checked for cleanliness before being allowed to the table; and although intended to keep terrestrial germs from space, this technology has awesome applications here on Mother Earth. The bacterial spore-detection unit can recognize anthrax and other harmful, spore-forming bacteria and alert people of the impending danger. As evidenced in the anthrax mailings of fall 2001 in the United States, the first sign of anthrax exposure was when people experienced flu-like symptoms, which unfortunately, can take as much as a week to develop after contamination. Anthrax cost 5 people their lives and infected 19 others; and the threat of bioterrorism became a routine concern, with new threats popping up nearly everyday. The attacks threatened the safety that so many Americans took for granted, as the very air that people breathed became suspect. Any building with a circulation system, where large groups congregate, was now a potential target.

Cytometric methods for measuring bacteria in water: advantages, pitfalls and applications.

PubMed

Hammes, Frederik; Egli, Thomas

2010-06-01

Rapid detection of microbial cells is a challenge in microbiology, particularly when complex indigenous communities or subpopulations varying in viability, activity and physiological state are investigated. Flow cytometry (FCM) has developed during the last 30 years into a multidisciplinary technique for analysing bacteria. When used correctly, FCM can provide a broad range of information at the single-cell level, including (but not limited to) total counts, size measurements, nucleic acid content, cell viability and activity, and detection of specific bacterial groups or species. The main advantage of FCM is that it is fast and easy to perform. It is a robust technique, which is adaptable to different types of samples and methods, and has much potential for automation. Hence, numerous FCM applications have emerged in industrial biotechnology, food and pharmaceutical quality control, routine monitoring of drinking water and wastewater systems, and microbial ecological research in soils and natural aquatic habitats. This review focuses on the information that can be gained from the analysis of bacteria in water, highlighting some of the main advantages, pitfalls and applications.

Influence of Soap Characteristics and Food Service Facility Type on the Degree of Bacterial Contamination of Open, Refillable Bulk Soaps.

PubMed

Schaffner, Donald W; Jensen, Dane; Gerba, Charles P; Shumaker, David; Arbogast, James W

2018-02-01

Concern has been raised regarding the public health risks from refillable bulk-soap dispensers because they provide an environment for potentially pathogenic bacteria to grow. This study surveyed the microbial quality of open refillable bulk soap in four different food establishment types in three states. Two hundred ninety-six samples of bulk soap were collected from food service establishments in Arizona, New Jersey, and Ohio. Samples were tested for total heterotrophic viable bacteria, Pseudomonas, coliforms and Escherichia coli, and Salmonella. Bacteria were screened for antibiotic resistance. The pH, solids content, and water activity of all soap samples were measured. Samples were assayed for the presence of the common antibacterial agents triclosan and parachlorometaxylenol. More than 85% of the soap samples tested contained no detectable microorganisms, but when a sample contained any detectable microorganisms, it was most likely contaminated at a very high level (∼7 log CFU/mL). Microorganisms detected in contaminated soap included Klebsiella oxytoca, Serratia liquefaciens, Shigella sonnei, Enterobacter gergoviae, Serratia odorifera, and Enterobacter cloacae. Twenty-three samples contained antibiotic-resistant organisms, some of which were resistant to two or more antibiotics. Every sample containing less than 4% solids had some detectable level of bacteria, whereas no samples with greater than 14% solids had detectable bacteria. This finding suggests the use of dilution and/or low-cost formulations as a cause of bacterial growth. There was a statistically significant difference ( P = 0.0035) between the fraction of bacteria-positive samples with no detected antimicrobial agent (17%) and those containing an antimicrobial agent (7%). Fast food operations and grocery stores were more likely to have detectable bacteria in bulk-soap samples compared with convenience stores ( P < 0.05). Our findings underscore the risk to public health from use of refillable bulk-soap dispensers in food service establishments.

Review on SERS of Bacteria

PubMed Central

Mosier-Boss, Pamela A.

2017-01-01

Surface enhanced Raman spectroscopy (SERS) has been widely used for chemical detection. Moreover, the inherent richness of the spectral data has made SERS attractive for use in detecting biological materials, including bacteria. This review discusses methods that have been used to obtain SERS spectra of bacteria. The kinds of SERS substrates employed to obtain SERS spectra are discussed as well as how bacteria interact with silver and gold nanoparticles. The roll of capping agents on Ag/Au NPs in obtaining SERS spectra is examined as well as the interpretation of the spectral data. PMID:29137201

Pseudomonas screening assay

NASA Technical Reports Server (NTRS)

Margalit, Ruth (Inventor)

1993-01-01

A method for the detection of Pseudomonas bacteria is described where an Azurin-specific antibody is employed for detecting the presence of Azurin in a test sample. The detection of the presence of Azurin in the sample is a conclusive indicator of the presence of the Pseudomonas bacteria since the Azurin protein is a specific marker for this bacterial strain.

9 CFR 113.25 - Culture media for detection of bacteria and fungi.

Code of Federal Regulations, 2013 CFR

2013-01-01

... 9 Animals and Animal Products 1 2013-01-01 2013-01-01 false Culture media for detection of... STANDARD REQUIREMENTS Standard Procedures § 113.25 Culture media for detection of bacteria and fungi. (a..., shall conform to such standards. In lieu of preparing the media from the individual ingredients, they...

9 CFR 113.25 - Culture media for detection of bacteria and fungi.

Code of Federal Regulations, 2012 CFR

2012-01-01

... 9 Animals and Animal Products 1 2012-01-01 2012-01-01 false Culture media for detection of... STANDARD REQUIREMENTS Standard Procedures § 113.25 Culture media for detection of bacteria and fungi. (a..., shall conform to such standards. In lieu of preparing the media from the individual ingredients, they...

9 CFR 113.25 - Culture media for detection of bacteria and fungi.

Code of Federal Regulations, 2014 CFR

2014-01-01

... 9 Animals and Animal Products 1 2014-01-01 2014-01-01 false Culture media for detection of... STANDARD REQUIREMENTS Standard Procedures § 113.25 Culture media for detection of bacteria and fungi. (a..., shall conform to such standards. In lieu of preparing the media from the individual ingredients, they...

Microfluidic electrochemical assay for rapid detection and quantification of Escherichia coli.

PubMed

Safavieh, Mohammadali; Ahmed, Minhaz Uddin; Tolba, Mona; Zourob, Mohammed

2012-01-15

Microfluidic electrochemical biosensor for performing Loop-mediated isothermal amplification (LAMP) was developed for the detection and quantification of Escherichia coli. The electrochemical detection for detecting the DNA amplification was achieved using Hoechst 33258 redox molecule and linear sweep voltametry (LSV). The DNA aggregation and minor groove binding with redox molecule cause a significant drop in the anodic oxidation of LSV. Unlike other electrochemical techniques, this method does not require the probe immobilization and the detection of the bacteria can be accomplished in a single chamber without DNA extraction and purification steps. The isothermal amplification time has a major role in the quantification of the bacteria. We have shown that we could detect and quantify 24 CFU/ml of bacteria and 8.6 fg/μl DNA in 60 min and 48 CFU/ml of bacteria in 35 min in LB media and urine samples. We believe that this microfluidic chip has great potential to be used as a point of care diagnostic (POC) device in the clinical/hospital application. Copyright © 2011 Elsevier B.V. All rights reserved.

Evaluation of Polymerase Chain Reaction for Detecting Coliform Bacteria in Drinking Water Sources

PubMed Central

Isfahani, Bahram Nasr; Fazeli, Hossein; Babaie, Zeinab; Poursina, Farkhondeh; Moghim, Sharareh; Rouzbahani, Meisam

2017-01-01

Background: Coliform bacteria are used as indicator organisms for detecting fecal pollution in water. Traditional methods including microbial culture tests in lactose-containing media and enzyme-based tests for the detection of β-galactosidase; however, these methods are time-consuming and less specific. The aim of this study was to evaluate polymerase chain reaction (PCR) for detecting coliform. Materials and Methods: Totally, 100 of water samples from Isfahan drinking water source were collected. Coliform bacteria and Escherichia coli were detected in drinking water using LacZ and LamB genes in PCR method performed in comparison with biochemical tests for all samples. Results: Using phenotyping, 80 coliform isolates were found. The results of the biochemical tests illustrated 78.7% coliform bacteria and 21.2% E. coli. PCR results for LacZ and LamB genes were 67.5% and 17.5%, respectively. Conclusion: The PCR method was shown to be an effective, sensitive, and rapid method for detecting coliform and E. coli in drinking water from the Isfahan drinking water sources. PMID:29142893

Surface-enhanced Raman scattering spectroscopy characterization and identification of foodborne bacteria

NASA Astrophysics Data System (ADS)

Liu, Yongliang; Chen, Yud-Ren; Nou, Xiangwu; Chao, Kaunglin

2007-09-01

Rapid and routine identification of foodborne bacteria are considerably important, because of bio- / agro- terrorism threats, public health concerns, and economic loss. Conventional, PCR, and immunoassay methods for the detection of bacteria are generally time-consuming, chemical reagent necessary and multi-step procedures. Fast microbial detection requires minimal sample preparation, permits the routine analysis of large numbers of samples with negligible reagent costs, and is easy to operate. Therefore, we have developed silver colloidal nanoparticle based surface-enhanced Raman scattering (SERS) spectroscopy as a potential tool for the rapid and routine detection of E. coli and L. monocytogenes. This study presents the further results of our examination on S. typhimonium, one of the most commonly outbreak bacteria, for the characteristic bands and subsequent identification.

Evaluation of a polymerase chain reaction-based system for detection of Salmonella enteritidis, Escherichia coli O157:H7, Listeria spp., and Listeria monocytogenes on fresh fruits and vegetables.

PubMed

Shearer, A E; Strapp, C M; Joerger, R D

2001-06-01

A polymerase chain reaction (PCR)-based detection system, BAX, was evaluated for its sensitivity in detecting Salmonella Enteritidis, Escherichia coli O157:H7, Listeria sp., and Listeria monocytogenes on fresh produce. Fifteen different types of produce (alfalfa sprouts, green peppers, parsley, white cabbage, radishes, onions, carrots, mushrooms, leaf lettuce, tomatoes, strawberries, cantaloupe, mango, apples, and oranges) were inoculated, in separate studies, with Salmonella Enteritidis, E. coli O157:H7, and L. monocytogenes down to the predicted level of 1 CFU per 25-g sample. Detection by BAX was compared to recovery of the inoculated bacteria by culture methods according to the Food and Drug Administration's (FDA) Bacteriological Analytical Manual (BAM). BAX was essentially as sensitive as the culture-based method in detecting Salmonella Enteritidis and L. monocytogenes and more sensitive than the culture-based method for the detection of E. coli O157:H7 on green pepper, carrot, radish, and sprout samples. Detection of the pathogenic bacteria in samples spiked with a predicted number of less than 10 CFU was possible for most produce samples, but both methods failed to detect L. monocytogenes on carrot samples and one of two mushroom and onion samples spiked with less than 100 CFU. Both BAX and the culture method were also unable to consistently recover low numbers of E. coli O157:H7 from alfalfa sprouts. The PCR method allowed detection of Salmonella Enteritidis, E. coli O157:H7, and L. monocytogenes at least 2 days earlier than the conventional culture methods.

Culture-free, highly sensitive, quantitative detection of bacteria from minimally processed samples using fluorescence imaging by smartphone.

PubMed

Shrivastava, Sajal; Lee, Won-Il; Lee, Nae-Eung

2018-06-30

A critical unmet need in the diagnosis of bacterial infections, which remain a major cause of human morbidity and mortality, is the detection of scarce bacterial pathogens in a variety of samples in a rapid and quantitative manner. Herein, we demonstrate smartphone-based detection of Staphylococcus aureus in a culture-free, rapid, quantitative manner from minimally processed liquid samples using aptamer-functionalized fluorescent magnetic nanoparticles. The tagged S. aureus cells were magnetically captured in a detection cassette, and then fluorescence was imaged using a smartphone camera with a light-emitting diode as the excitation source. Our results showed quantitative detection capability with a minimum detectable concentration as low as 10 cfu/ml by counting individual bacteria cells, efficiently capturing S. aureus cells directly from a peanut milk sample within 10 min. When the selectivity of detection was investigated using samples spiked with other pathogenic bacteria, no significant non-specific detection occurred. Furthermore, strains of S. aureus from various origins showed comparable results, ensuring that the approach can be widely adopted. Therefore, the quantitative fluorescence imaging platform on a smartphone could allow on-site detection of bacteria, providing great potential assistance during major infectious disease outbreaks in remote and resource-limited settings. Copyright © 2018 Elsevier B.V. All rights reserved.

ETV Tech Brief: Rapid Fungi and Bacteria Detection Technologies

EPA Science Inventory

Technical brief that summarizes the results for Mycometer, Inc. Mycometer®-test and Bactiquant®-test, which are rapid detection technologies for fungi and bacteria. The brief summarizes the results of the verification report and statement.

Biosensors for Whole-Cell Bacterial Detection

PubMed Central

Rushworth, Jo V.; Hirst, Natalie A.; Millner, Paul A.

2014-01-01

SUMMARY Bacterial pathogens are important targets for detection and identification in medicine, food safety, public health, and security. Bacterial infection is a common cause of morbidity and mortality worldwide. In spite of the availability of antibiotics, these infections are often misdiagnosed or there is an unacceptable delay in diagnosis. Current methods of bacterial detection rely upon laboratory-based techniques such as cell culture, microscopic analysis, and biochemical assays. These procedures are time-consuming and costly and require specialist equipment and trained users. Portable stand-alone biosensors can facilitate rapid detection and diagnosis at the point of care. Biosensors will be particularly useful where a clear diagnosis informs treatment, in critical illness (e.g., meningitis) or to prevent further disease spread (e.g., in case of food-borne pathogens or sexually transmitted diseases). Detection of bacteria is also becoming increasingly important in antibioterrorism measures (e.g., anthrax detection). In this review, we discuss recent progress in the use of biosensors for the detection of whole bacterial cells for sensitive and earlier identification of bacteria without the need for sample processing. There is a particular focus on electrochemical biosensors, especially impedance-based systems, as these present key advantages in terms of ease of miniaturization, lack of reagents, sensitivity, and low cost. PMID:24982325

Chlorophyll a might structure a community of potentially pathogenic culturable Vibrionaceae. Insights from a one-year study of water and mussels surveyed on the French Atlantic coast.

PubMed

Deter, J; Lozach, S; Derrien, A; Véron, A; Chollet, J; Hervio-Heath, D

2010-02-01

The present study focused on the isolation of culturable bacteria from mussels and sea water to identify Vibrionaceae potentially pathogenic for humans. Three sites located on the French Atlantic coast were monitored monthly (twice each month during summer) for 1 year. Environmental parameters were surveyed (water temperature, salinity, turbidity, chlorophyll a) and bacteria were detected by culture and identified by API 20E(®) systems (BioMérieux) and PCR. A total of seven species were detected (Grimontia hollisae, Photobacterium damselae, Vibrio alginolyticus, V. cholerae, V. fluvialis, V. vulnificus and V. parahaemolyticus) and species diversity was higher at the end of summer. Surprisingly, V. cholerae non-O1/non-O139 was detected in spring. No site effect was detected. Using Sørensen similarity indices and statistical analyses, we showed that chlorophyll a had a significant influence on the bacterial community detected in mussels and assemblages were more similar to one another when chlorophyll a values were above 20 µg l(-1) . No significant effect of any parameter was found on the community detected in water samples. Such surveys are essential for the understanding of sanitary crises and detection of emerging pathogens. © 2010 Society for Applied Microbiology and Blackwell Publishing Ltd.

Quantification and identification of particle-associated bacteria in unchlorinated drinking water from three treatment plants by cultivation-independent methods.

PubMed

Liu, G; Ling, F Q; Magic-Knezev, A; Liu, W T; Verberk, J Q J C; Van Dijk, J C

2013-06-15

Water quality regulations commonly place quantitative limits on the number of organisms (e.g., heterotrophic plate count and coliforms) without considering the presence of multiple cells per particle, which is only counted as one regardless how many cells attached. Therefore, it is important to quantify particle-associated bacteria (PAB), especially cells per particle. In addition, PAB may house (opportunistic) pathogens and have higher resistance to disinfection than planktonic bacteria. It is essential to know bacterial distribution on particles. However, limited information is available on quantification and identification of PAB in drinking water. In the present study, PAB were sampled from the unchlorinated drinking water at three treatment plants in the Netherlands, each with different particle compositions. Adenosine triphosphate (ATP) and total cell counts (TCC) with flow cytometry were used to quantify the PAB, and high-throughput pyrosequencing was used to identify them. The number and activity of PAB ranged from 1.0 to 3.5 × 10(3) cells ml(-1) and 0.04-0.154 ng l(-1) ATP. There were between 25 and 50 cells found to be attached on a single particle. ATP per cell in PAB was higher than in planktonic bacteria. Among the identified sequences, Proteobacteria were found to be the most dominant phylum at all locations, followed by OP3 candidate division and Nitrospirae. Sequences related to anoxic bacteria from the OP3 candidate division and other anaerobic bacteria were detected. Genera of bacteria were found appear to be consistent with the major element composition of the associated particles. The presence of multiple cells per particle challenges the use of quantitative methods such as HPC and Coliforms that are used in the current drinking water quality regulations. The detection of anoxic and anaerobic bacteria suggests the ecological importance of PAB in drinking water distribution systems. Copyright © 2013 Elsevier Ltd. All rights reserved.

Detection and discrimination of microorganisms on various substrates with quantum cascade laser spectroscopy

NASA Astrophysics Data System (ADS)

Padilla-Jiménez, Amira C.; Ortiz-Rivera, William; Rios-Velazquez, Carlos; Vazquez-Ayala, Iris; Hernández-Rivera, Samuel P.

2014-06-01

Investigations focusing on devising rapid and accurate methods for developing signatures for microorganisms that could be used as biological warfare agents' detection, identification, and discrimination have recently increased significantly. Quantum cascade laser (QCL)-based spectroscopic systems have revolutionized many areas of defense and security including this area of research. In this contribution, infrared spectroscopy detection based on QCL was used to obtain the mid-infrared (MIR) spectral signatures of Bacillus thuringiensis, Escherichia coli, and Staphylococcus epidermidis. These bacteria were used as microorganisms that simulate biothreats (biosimulants) very truthfully. The experiments were conducted in reflection mode with biosimulants deposited on various substrates including cardboard, glass, travel bags, wood, and stainless steel. Chemometrics multivariate statistical routines, such as principal component analysis regression and partial least squares coupled to discriminant analysis, were used to analyze the MIR spectra. Overall, the investigated infrared vibrational techniques were useful for detecting target microorganisms on the studied substrates, and the multivariate data analysis techniques proved to be very efficient for classifying the bacteria and discriminating them in the presence of highly IR-interfering media.

Detection of Viral and Bacterial Pathogens in Hospitalized Children With Acute Respiratory Illnesses, Chongqing, 2009–2013

PubMed Central

Wei, Lan; Liu, Wei; Zhang, Xiao-Ai; Liu, En-Mei; Wo, Yin; Cowling, Benjamin J.; Cao, Wu-Chun

2015-01-01

Abstract Acute respiratory infections (ARIs) cause large disease burden each year. The codetection of viral and bacterial pathogens is quite common; however, the significance for clinical severity remains controversial. We aimed to identify viruses and bacteria in hospitalized children with ARI and the impact of mixed detections. Hospitalized children with ARI aged ≤16 were recruited from 2009 to 2013 at the Children's Hospital of Chongqing Medical University, Chongqing, China. Nasopharyngeal aspirates (NPAs) were collected for detection of common respiratory viruses by reverse transcription polymerase chain reaction (RT-PCR) or PCR. Bacteria were isolated from NPAs by routine culture methods. Detection and codetection frequencies and clinical features and severity were compared. Of the 3181 hospitalized children, 2375 (74.7%) were detected with ≥1 virus and 707 (22.2%) with ≥1 bacteria, 901 (28.3%) with ≥2 viruses, 57 (1.8%) with ≥2 bacteria, and 542 (17.0%) with both virus and bacteria. The most frequently detected were Streptococcus pneumoniae, respiratory syncytial virus, parainfluenza virus, and influenza virus. Clinical characteristics were similar among different pathogen infections for older group (≥6 years old), with some significant difference for the younger. Cases with any codetection were more likely to present with fever; those with ≥2 virus detections had higher prevalence of cough; cases with virus and bacteria codetection were more likely to have cough and sputum. No significant difference in the risk of pneumonia, severe pneumonia, and intensive care unit admission were found for any codetection than monodetection. There was a high codetection rate of common respiratory pathogens among hospitalized pediatric ARI cases, with fever as a significant predictor. Cases with codetection showed no significant difference in severity than those with single pathogens. PMID:25906103

Cyclic Dinucleotides in Oral Bacteria and in Oral Biofilms.

PubMed

Gürsoy, Ulvi K; Gürsoy, Mervi; Könönen, Eija; Sintim, Herman O

2017-01-01

Oral cavity acts as a reservoir of bacterial pathogens for systemic infections and several oral microorganisms have been linked to systemic diseases. Quorum sensing and cyclic dinucleotides, two "decision-making" signaling systems, communicate to regulate physiological process in bacteria. Discovery of cyclic dinucleotides has a long history, but the progress in our understanding of how cyclic dinucleotides regulate bacterial lifestyle is relatively new. Oral microorganisms form some of the most intricate biofilms, yet c-di-GMP, and c-di-AMP signaling have been rarely studied in oral biofilms. Recent studies demonstrated that, with the aid of bacterial messenger molecules and their analogs, it is possible to activate host innate and adaptive immune responses and epithelial integrity with a dose that is relevant to inhibit bacterial virulence mechanisms, such as fimbriae and exopolysaccharide production, biofilm formation, and host cell invasion. The aim of this perspective article is to present available information on cyclic dinucleotides in oral bacteria and in oral biofilms. Moreover, technologies that can be used to detect cyclic dinucleotides in oral biofilms are described. Finally, directions for future research are highlighted.

Comparative evaluation of Oxoid Signal and BACTEC radiometric blood culture systems for the detection of bacteremia and fungemia

DOE Office of Scientific and Technical Information (OSTI.GOV)

Weinstein, M.P.; Mirrett, S.; Reller, L.B.

1988-05-01

The Oxoid Signal blood culture system is a newly described, innovative method for visually detecting growth of microorganisms. We did 5,999 paired comparisons of equal volumes (10 ml) of blood in the Oxoid Signal and BACTEC radiometric blood culture systems at two university hospitals that use identical methods of obtaining and processing specimens. Overall, more microorganisms were detected in the BACTEC system (P less than 0.001), in particular, streptococci (P less than 0.01), fungi (P less than 0.001), and nonfermentative gram-negative rods, especially Acinetobacter species (P less than 0.001). Trends favoring the BACTEC system for detection of Pseudomonas aeruginosa, Haemophilusmore » species, and Neisseria species were noted. There were no differences in the yield of staphylococci, members of the family Enterobacteriaceae, and anaerobic bacteria. When both systems detected sepsis, the BACTEC did so earlier (P less than 0.001). This advantage was most notable at 24 h (70% of BACTEC positives detected versus 48% of Oxoid positives). The proportion of positives detected after 48 h, however, was similar (BACTEC, 84%; Oxoid, 78%). Revisions in the Oxoid Signal system itself or in the processing of Oxoid bottles appear to be necessary to improve its performance in detecting certain microorganism groups, especially fungi.« less

Label-Free in Situ Discrimination of Live and Dead Bacteria by Surface-Enhanced Raman Scattering.

PubMed

Zhou, Haibo; Yang, Danting; Ivleva, Natalia P; Mircescu, Nicoleta E; Schubert, Sören; Niessner, Reinhard; Wieser, Andreas; Haisch, Christoph

2015-07-07

Techniques to distinguish between live and dead bacteria in a quantitative manner are in high demand in numerous fields including medical care, food safety, and public security as well as basic science research. This work demonstrates new nanostructures (silver nanoparticles coating bacteria structure, Bacteria@AgNPs) and their utility for rapid counting of live and dead bacteria by surface-enhanced Raman scattering (SERS). We found that suspensions containing Gram-negative organisms as well as AgNPs give strong SERS signals of live bacteria when generated selectively on the particle surface. However, almost no SERS signals can be detected from Bacteria@AgNPs suspensions containing dead bacteria. We demonstrate successful quantification of different percentages of dead bacteria both in bulk liquid and on glass surfaces by using SERS mapping on a single cell basis. Furthermore, different chemicals have been used to elucidate the mechanism involved in this observation. Finally, we used the Bacteria@AgNPs method to detect antibiotic resistance of E. coli strains against several antibiotics used in human medicine.

Development of bacteria-based bioassays for arsenic detection in natural waters.

PubMed

Diesel, Elizabeth; Schreiber, Madeline; van der Meer, Jan Roelof

2009-06-01

Arsenic contamination of natural waters is a worldwide concern, as the drinking water supplies for large populations can have high concentrations of arsenic. Traditional techniques to detect arsenic in natural water samples can be costly and time-consuming; therefore, robust and inexpensive methods to detect arsenic in water are highly desirable. Additionally, methods for detecting arsenic in the field have been greatly sought after. This article focuses on the use of bacteria-based assays as an emerging method that is both robust and inexpensive for the detection of arsenic in groundwater both in the field and in the laboratory. The arsenic detection elements in bacteria-based bioassays are biosensor-reporter strains; genetically modified strains of, e.g., Escherichia coli, Bacillus subtilis, Staphylococcus aureus, and Rhodopseudomonas palustris. In response to the presence of arsenic, such bacteria produce a reporter protein, the amount or activity of which is measured in the bioassay. Some of these bacterial biosensor-reporters have been successfully utilized for comparative in-field analyses through the use of simple solution-based assays, but future methods may concentrate on miniaturization using fiberoptics or microfluidics platforms. Additionally, there are other potential emerging bioassays for the detection of arsenic in natural waters including nematodes and clams.

Detecting bacteria and Determining Their Susceptibility to Antibiotics by Stochastic Confinement in Nanoliter Droplets using Plug-Based Microfluidics

DOE Office of Scientific and Technical Information (OSTI.GOV)

Boedicker, J.; Li, L; Kline, T

2008-01-01

This article describes plug-based microfluidic technology that enables rapid detection and drug susceptibility screening of bacteria in samples, including complex biological matrices, without pre-incubation. Unlike conventional bacterial culture and detection methods, which rely on incubation of a sample to increase the concentration of bacteria to detectable levels, this method confines individual bacteria into droplets nanoliters in volume. When single cells are confined into plugs of small volume such that the loading is less than one bacterium per plug, the detection time is proportional to plug volume. Confinement increases cell density and allows released molecules to accumulate around the cell, eliminatingmore » the pre-incubation step and reducing the time required to detect the bacteria. We refer to this approach as stochastic confinement. Using the microfluidic hybrid method, this technology was used to determine the antibiogram - or chart of antibiotic sensitivity - of methicillin-resistant Staphylococcus aureus (MRSA) to many antibiotics in a single experiment and to measure the minimal inhibitory concentration (MIC) of the drug cefoxitin (CFX) against this strain. In addition, this technology was used to distinguish between sensitive and resistant strains of S. aureus in samples of human blood plasma. High-throughput microfluidic techniques combined with single-cell measurements also enable multiple tests to be performed simultaneously on a single sample containing bacteria. This technology may provide a method of rapid and effective patient-specific treatment of bacterial infections and could be extended to a variety of applications that require multiple functional tests of bacterial samples on reduced timescales.« less

Method and apparatus for detecting phycocyanin-pigmented algae and bacteria from reflected light

NASA Technical Reports Server (NTRS)

Vincent, Robert (Inventor)

2013-01-01

The present invention relates to a method of detecting phycocyanin algae or bacteria in water from reflected light, and also includes devices for the measurement, calculation and transmission of data relating to that method.

Method and apparatus for detecting phycocyanin-pigmented algae and bacteria from reflected light

NASA Technical Reports Server (NTRS)

Vincent, Robert (Inventor)

2006-01-01

The present invention relates to a method of detecting phycocyanin algae or bacteria in water from reflected light, and also includes devices for the measurement, calculation and transmission of data relating to that method.

New medium for detection of esterase and gelatinase activity.

PubMed

Pácová, Z; Kocur, M

1984-10-01

A new medium was developed for detecting esterase and gelatinase activities in aerobic and facultatively anaerobic bacteria. The new medium was tested with various strains of bacteria and the results showed agreement between the reactions in the new medium and those obtained by conventional techniques. The new medium is more economical and may be used for a rapid differentiation of Serratia, Aeromonas and Vibrio species from biochemically similar bacteria.

PCR method for the rapid detection and discrimination of Legionella spp. based on the amplification of pcs, pmtA, and 16S rRNA genes.

PubMed

Janczarek, Monika; Palusińska-Szysz, Marta

2016-05-01

Legionella bacteria are organisms of public health interest due to their ability to cause pneumonia (Legionnaires' disease) in susceptible humans and their ubiquitous presence in water supply systems. Rapid diagnosis of Legionnaires' disease allows the use of therapy specific for the disease. L. pneumophila serogroup 1 is the most common cause of infection acquired in community and hospital environments. The non-L. pneumophila infections are likely under-detected because of a lack of effective diagnosis. In this work, simplex and duplex PCR assays with the use of new molecular markers pcs and pmtA involved in phosphatidylcholine synthesis were specified for rapid and cost-efficient identification and distinguishing Legionella species. The sets of primers developed were found to be sensitive and specific for reliable detection of Legionella belonging to the eight most clinically relevant species. Among these, four primer sets I, II, VI, and VII used for duplex-PCRs proved to have the highest identification power and reliability in the detection of the bacteria. Application of this PCR-based method should improve detection of Legionella spp. in both clinical and environmental settings and facilitate molecular typing of these organisms.

Persistence of bacterial DNA in orthopedic infections.

PubMed

Kaplan, Heidi B; Miranda, Justin A; Gogola, Gloria R; Gomez, Karen; Ambrose, Catherine G

2018-06-01

Polymerase chain reaction (PCR) has been proposed as a method to identify bacteria in clinical samples because it is more sensitive than culture techniques and can produce results rapidly. However, PCR can detect DNA from dead cells and thus cannot distinguish between live and dead cells in a tissue sample. Killed Staphylococcus aureus cells were implanted into the femurs and knee joints of rats to determine the length of time that DNA from dead cells is detectable in a living animal under conditions similar to common orthopedic infections. In the joint infection model studied here, the DNA from the dead planktonic bacteria was detected using PCR immediately after injection or 24 h later, but was undetectable 48 and 72 h after injection. In the biofilm implanted-device model studied, the DNA from these dead biofilm cells was detected by PCR immediately after implantation and at 24 h, but not at 48 or 72 h. Thus, our results indicate that DNA from dead cells does not persist in these animal model systems for more than 2 days, which should reduce concerns about possible false positive results using molecular DNA-based techniques for the detection of pathogens. Copyright © 2018. Published by Elsevier Inc.

Magneto-actuated immunoassay for the detection of Mycobacterium fortuitum in hemodialysis water.

PubMed

Brugnera, Michelle Fernanda; Bundalian, Reynaldo; Laube, Tamara; Julián, Esther; Luquin, Marina; Zanoni, Maria Valnice Boldrin; Pividori, Maria Isabel

2016-06-01

This paper addresses a sensitive method for the detection of mycobacteria in hemodialysis water samples based on a magneto-actuated immunoassay with optical readout. In this approach, micro (2.8μm) sized magnetic particles were modified with an antibody against the lipoarabinomannan (LAM) located in the mycobacterial cell wall. The system relies on the immunocapturing of the mycobacteria with the tailored antiLAM magnetic particles to pre-concentrate the bacteria from the hemodialysis samples throughout an immunological reaction. The performance of the immunomagnetic separation on the magnetic carrier was evaluated using confocal microscopy to study the binding pattern, as well as a magneto-actuated immunoassay with optical readout for the rapid detection of the bacteria in spiked hemodialysis samples. In this approach, the antiLAM polyclonal antibody was labeled with fluorescein isothiocyanate. The optical readout was achieved by the incubation with a secondary anti-fluorescein antibody labeled with peroxidase as optical reporter. The magneto-actuated immunoassay was able to detect mycobacteria contamination in hemodialysis water at a limit of detection of 13CFUmL(-1) in a total assay time of 3h without any previous culturing pre-enrichment step. Copyright © 2016 Elsevier B.V. All rights reserved.

Detection of tlh and tdh genes in Vibrio Parahaemolyticus inhabiting farmed water ecosystem used for L. Vannamei aquaculture

NASA Astrophysics Data System (ADS)

Nawan Hasrimi, Adila; Budiharjo, Anto; Nur Jannah, Siti

2018-05-01

Vibrio parahaemolyticus is hallophilic gram-negative bacteria that live as natural inhabitant in aquatic environment. All Vibrio parahaemolyticus strain known to have thermolabile hemolysin encoded by tlh gene as species marker. Thermostable direct hemolysin encoded by tdh gene is responsible for regulating virulence factor in Vibrio parhaemolyticus. Aim of this research is to detect tlh and tdh gene from water of L. vannamei aquaculture in Rembang regency. Colonies of green-blueish bacteria grew from isolation of L. vannamei aquaculture water in CD-VP media which was identified as Vibrio parahaemolyticus. Colonies of V. parahaemolyticus grew to be small and green-blueish bacteria colonies in TCBS agar. Result of molecular analysis showed that bacteria isolated from water sample are specifically identified as Vibrio parahaemolyticus bacteria by the detection of tlh gene. Vibrio parahaemolyticus isolated from water of L. vannamei aquaculture detected as tdh negative that indicates tdh gene is not present in isolated bacteria. Vibrio parahaemolyticus isolate were cultured in Wagatsuma agar for tdh gene confirmation test that showed Kanagawa negative result, which indicated that V. parahaemolyticus did not produce thermostable direct hemolysin. These results showed that Vibrio parahaemolyticus isolated from aquatic environment of L. vannamei aquaculture in Rembang regency did not show virulence factors.

Enzymatic Digestion for Improved Bacteria Separation from Leafy Green Vegetables.

PubMed

Wang, Danhui; Wang, Ziyuan; He, Fei; Kinchla, Amanda J; Nugen, Sam R

2016-08-01

An effective and rapid method for the separation of bacteria from food matrix remains a bottleneck for rapid bacteria detection for food safety. Bacteria can strongly attach to a food surface or internalize within the matrix, making their isolation extremely difficult. Traditional methods of separating bacteria from food routinely involve stomaching, blending, and shaking. However, these methods may not be efficient at removing all the bacteria from complex matrices. Here, we investigate the benefits of using enzyme digestion followed by immunomagnetic separation to isolate Salmonella from spinach and lettuce. Enzymatic digestion using pectinase and cellulase was able to break down the structure of the leafy green vegetables, resulting in the detachment and release of Salmonella from the leaves. Immunomagnetic separation of Salmonella from the liquefied sample allowed an additional separation step to achieve a more pure sample without leaf debris that may benefit additional downstream applications. We have investigated the optimal combination of pectinase and cellulase for the digestion of spinach and lettuce to improve sample detection yields. The concentrations of enzymes used to digest the leaves were confirmed to have no significant effect on the viability of the inoculated Salmonella. Results reported that the recovery of the Salmonella from the produce after enzyme digestion of the leaves was significantly higher (P < 0.05) than traditional sample preparation methods to separate bacteria (stomaching and manually shaking). The results demonstrate the potential for use of enzyme digestion prior to separation can improve the efficiency of bacteria separation and increase the likelihood of detecting pathogens in the final detection assay.

DIYA: A Bacterial Annotation Pipeline for any Genomics Lab

DTIC Science & Technology

2009-02-12

make more modules available. Functions we are looking to add to the DIYA pipeline include software for detection of prophages, CRISPR elements (Sorek...R. et al. (2008) CRISPR –a widespread system that provides acquired resistance against phages in bacteria and archaea. Nat. Rev. Microbiol., 6, 181

Dielectrophoresis and dielectrophoretic impedance detection of adenovirus and rotavirus

NASA Astrophysics Data System (ADS)

Nakano, Michihiko; Ding, Zhenhao; Suehiro, Junya

2016-01-01

The aim of this study is the electrical detection of pathogenic viruses, namely, adenovirus and rotavirus, using dielectrophoretic impedance measurement (DEPIM). DEPIM consists of two simultaneous processes: dielectrophoretic trapping of the target and measurement of the impedance change and increase in conductance with the number of trapped targets. This is the first study of applying DEPIM, which was originally developed to detect bacteria suspended in aqueous solutions, to virus detection. The dielectric properties of the viruses were also investigated in terms of their dielectrophoretic behavior. Although their estimated dielectric properties were different from those of bacteria, the trapped viruses increased the conductance of the microelectrode in a manner similar to that in bacteria detection. We demonstrated the electrical detection of viruses within 60 s at concentrations as low as 70 ng/ml for adenovirus and 50 ng/ml for rotavirus.

Bacterial Landscape of Bloodstream Infections in Neutropenic Patients via High Throughput Sequencing

PubMed Central

Gyarmati, Peter; Kalin, Mats; Öhrmalm, Lars; Giske, Christian G.

2015-01-01

Background Bloodstream infection (BSI) is a common and potentially life-threatening complication in patients with hematological malignancies and therapy-induced neutropenia. Administration of broad spectrum antibiotics has substantially decreased the mortality rate in febrile neutropenia, but bacterial infection is documented in only one-third or fewer of the cases. BSI is typically diagnosed by blood culture; however, this method can detect only culturable pathogens. Methods In the present study, a total of 130 blood samples from hematological patients receiving dose-intensive antitumoural treatment were subjected to 16S rRNA PCR and 62 of them were cultured. PCR positive samples were processed to high throughput sequencing by amplifying the V1-V3 regions of the 16S rRNA gene to obtain a full spectrum of bacteria present in BSI. Results Five phyla and 30 genera were identified with sequencing compared to 2 phyla and 4 genera with culture. The largest proportion of bacteria detected by sequencing belonged to Proteobacteria (55.2%), Firmicutes (33.4%) and Actinobacteria (8.6%), while Fusobacteria (0.4%) and Bacteroidetes (0.1%) were also detected. Ninety-eight percent of the bacteria identified by sequencing were opportunistic human pathogens and 65% belonged to the normal human microbiota. Conclusions The present study indicates that BSIs in neutropenic hosts contain a much broader diversity of bacteria, likely with host origin, than previously realized. The elevated ratio of Proteobacteria in BSI corroborates the results found in other systemic inflammatory diseases, such as inflammatory bowel disease or mucosal infections. This knowledge may become of value for tailoring antimicrobial drug administration. PMID:26270467

Bacterial autolysins trim cell surface peptidoglycan to prevent detection by the Drosophila innate immune system

PubMed Central

Atilano, Magda Luciana; Pereira, Pedro Matos; Vaz, Filipa; Catalão, Maria João; Reed, Patricia; Grilo, Inês Ramos; Sobral, Rita Gonçalves; Ligoxygakis, Petros; Pinho, Mariana Gomes; Filipe, Sérgio Raposo

2014-01-01

Bacteria have to avoid recognition by the host immune system in order to establish a successful infection. Peptidoglycan, the principal constituent of virtually all bacterial surfaces, is a specific molecular signature recognized by dedicated host receptors, present in animals and plants, which trigger an immune response. Here we report that autolysins from Gram-positive pathogenic bacteria, enzymes capable of hydrolyzing peptidoglycan, have a major role in concealing this inflammatory molecule from Drosophila peptidoglycan recognition proteins (PGRPs). We show that autolysins trim the outermost peptidoglycan fragments and that in their absence bacterial virulence is impaired, as PGRPs can directly recognize leftover peptidoglycan extending beyond the external layers of bacterial proteins and polysaccharides. The activity of autolysins is not restricted to the producer cells but can also alter the surface of neighboring bacteria, facilitating the survival of the entire population in the infected host. DOI: http://dx.doi.org/10.7554/eLife.02277.001 PMID:24692449

Dual-Recognition Förster Resonance Energy Transfer Based Platform for One-Step Sensitive Detection of Pathogenic Bacteria Using Fluorescent Vancomycin-Gold Nanoclusters and Aptamer-Gold Nanoparticles.

PubMed

Yu, Mengqun; Wang, Hong; Fu, Fei; Li, Linyao; Li, Jing; Li, Gan; Song, Yang; Swihart, Mark T; Song, Erqun

2017-04-04

The effective monitoring, identification, and quantification of pathogenic bacteria is essential for addressing serious public health issues. In this study, we present a universal and facile one-step strategy for sensitive and selective detection of pathogenic bacteria using a dual-molecular affinity-based Förster (fluorescence) resonance energy transfer (FRET) platform based on the recognition of bacterial cell walls by antibiotic and aptamer molecules, respectively. As a proof of concept, Vancomycin (Van) and a nucleic acid aptamer were employed in a model dual-recognition scheme for detecting Staphylococcus aureus (Staph. aureus). Within 30 min, by using Van-functionalized gold nanoclusters and aptamer-modified gold nanoparticles as the energy donor and acceptor, respectively, the FRET signal shows a linear variation with the concentration of Staph. aureus in the range from 20 to 10 8 cfu/mL with a detection limit of 10 cfu/mL. Other nontarget bacteria showed negative results, demonstrating the good specificity of the approach. When employed to assay Staph. aureus in real samples, the dual-recognition FRET strategy showed recoveries from 99.00% to the 109.75% with relative standard derivations (RSDs) less than 4%. This establishes a universal detection platform for sensitive, specific, and simple pathogenic bacteria detection, which could have great impact in the fields of food/public safety monitoring and infectious disease diagnosis.

Detection of bacteria in suspension using a superconducting Quantum interference device

DOE Office of Scientific and Technical Information (OSTI.GOV)

Grossman, H.L.; Myers, W.R.; Vreeland, V.J.

2003-06-09

We demonstrate a technique for detecting magnetically-labeled Listeria monocytogenes and for measuring the binding rate between antibody-linked magnetic particles and bacteria. This assay, which is both sensitive and straightforward to perform, can quantify specific bacteria in a sample without the need to immobilize the bacteria or wash away unbound magnetic particles. In the measurement, we add 50 nm diameter superparamagnetic particles, coated with antibodies, to a liquid sample containing L. monocytogenes. We apply a pulsed magnetic field to align the magnetic dipole moments and use a high transition temperature Superconducting Quantum Interference Device (SQUID), an extremely sensitive detector of magneticmore » flux, to measure the magnetic relaxation signal when the field is turned off. Unbound particles randomize direction by Brownian rotation too quickly to be detected. In contrast, particles bound to L. monocytogenes are effectively immobilized and relax in about 1 s by rotation of the internal dipole moment. This Neel relaxation process is detected by the SQUID. The measurements indicate a detection limit of (5.6 {+-} 1.1) x 10{sup 6} L. monocytogenes for a 20 {micro}L sample volume. If the sample volume were reduced to 1 nL, we estimate that the detection limit could be improved to 230 {+-} 40 L. monocytogenes cells. Time-resolved measurements yield the binding rate between the particles and bacteria.« less

Design and field application of a UV-LED based optical fiber biofilm sensor.

PubMed

Fischer, Matthias; Wahl, Martin; Friedrichs, Gernot

2012-03-15

Detecting changes in the formation dynamics of biofilms stemming from bacteria and unicellular microorganisms in their natural environment is of prime interest for biological, ecological as well as anti-fouling technology research. We developed a robust optical fiber-based biofilm sensor ready to be applied in natural aquatic environments for on-line, in situ and non-destructive monitoring of large-area biofilms. The device is based on the detection of the natural fluorescence of microorganisms constituting the biofilm. Basically, the intrinsic fluorescence of the amino acid tryptophan is excited at a wavelength of λ=280 nm and detected at λ=350 nm utilising a numerically optimized sensor head equipped with a UV-LED light source and optical fiber bundles for efficient fluorescence light collection. Calibration was carried out with tryptophan solutions and two characteristic marine bacteria strains revealing linear signal response, satisfactory background suppression, wide dynamic range, and an experimental detection limit of 4 × 10(3)cells/cm(2). Successful field experiments in the Baltic Sea accomplished over a period of twenty-one days provided for the first time continuous observation of biofilm formation dynamics in a natural habitat. Starting from the first adhering bacteria, the measurement yielded the characteristic three phases of biofilm formation up to a fully developed biofilm. The sensor system holds potential for applications in aquatic sciences including deep sea research and, after further miniaturisation, in the industrial and biomedical field. Copyright © 2012 Elsevier B.V. All rights reserved.

Comparison of bacteriological culture and PCR for detection of bacteria in ovine milk--sheep are not small cows.

PubMed

Zadoks, Ruth N; Tassi, Riccardo; Martin, Elena; Holopainen, Jani; McCallum, Sarah; Gibbons, James; Ballingall, Keith T

2014-10-01

Mastitis, inflammation of the mammary gland, is an important cause of disease, mortality, and production losses in dairy and meat sheep. Mastitis is commonly caused by intramammary infection with bacteria, which can be detected by bacterial culture or PCR. PathoProof (Thermo Fisher Scientific Ltd., Vantaa, Finland) is a commercially available real-time PCR system for the detection of bovine mastitis pathogens. Sheep differ from cattle in the bacterial species or bacterial strains that cause mastitis, as well as in the composition of their milk. The aim of this study was to evaluate whether the PathoProof system was suitable for detection of mastitis pathogens in sheep milk. Milk samples were collected aseptically from 219 udder halves of 113 clinically healthy ewes in a single flock. Aliquots were used for bacteriological culture and real-time PCR-based detection of bacteria. For species identified by culture, the diagnosis was confirmed by species-specific conventional PCR or by sequencing of a housekeeping gene. The majority of samples were negative by culture (74.4% of 219 samples) and real-time PCR (82.3% of 192 samples). Agreement was observed for 138 of 192 samples. Thirty-four samples were positive by culture only, mostly due to presence of species that are not covered by primers in the PCR system (e.g., Mannheimia spp.). Two samples were positive for Streptococcus uberis by culture but not by PCR directly from the milk samples. This was not due to inability of the PCR primers to amplify ovine Streptococcus uberis, as diluted DNA extracts from the same samples and DNA extracts from the bacterial isolates were positive by real-time PCR. For samples containing Staphylococcus spp., 11 samples were positive by culture and PCR, 9 by culture only, and 20 by PCR only. Samples that were negative by either method had lower bacterial load than samples that were positive for both methods, whereas no clear relation with species identity was observed. This study provides proof of principle that real-time PCR can be used for detection of mastitis pathogens in ovine milk. Routine use in sheep may require inclusion of primer sets for sheep-specific mastitis pathogens. Copyright © 2014 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

Nasal septal abscess caused by anaerobic bacteria of oral flora.

PubMed

Hyo, Yukiyoshi; Fukushima, Hisaki; Harada, Tamotsu; Hara, Hirotaka

2018-06-07

Although nasal septal abscess (NSA) was formerly common, it has become rare since the development of antibiotics. NSA, if left untreated, can lead to intracranial complications such as meningitis and eventually result in saddle-nose deformity. NSA often occurs after injury, and indigenous skin bacteria such as Staphylococcus aureus are frequently detected. We treated a patient who had injured the upper alveolus in a fall on the stairs and developed NSA two weeks later. Anaerobic bacteria, including Veillonella parvula and Peptostreptococcus sp., were detected. Symptoms were relieved by needle and incisional drainage. Our patient represents a very rare case of NSA in terms of the cause of onset and the detected bacteria. Early drainage can result in good outcomes. Copyright © 2018 Elsevier B.V. All rights reserved.

9 CFR 113.27 - Detection of extraneous viable bacteria and fungi in live vaccines.

Code of Federal Regulations, 2010 CFR

2010-01-01

... bacteria and fungi in live vaccines. 113.27 Section 113.27 Animals and Animal Products ANIMAL AND PLANT... bacteria and fungi in live vaccines. Unless otherwise specified by the Administrator or elsewhere exempted... Seed Bacteria shall be tested for extraneous viable bacteria and fungi as prescribed in this section. A...

9 CFR 113.27 - Detection of extraneous viable bacteria and fungi in live vaccines.

Code of Federal Regulations, 2011 CFR

2011-01-01

... bacteria and fungi in live vaccines. 113.27 Section 113.27 Animals and Animal Products ANIMAL AND PLANT... bacteria and fungi in live vaccines. Unless otherwise specified by the Administrator or elsewhere exempted... Seed Bacteria shall be tested for extraneous viable bacteria and fungi as prescribed in this section. A...

Inactivation and injury of total coliform bacteria after primary disinfection of drinking water by TiO2 photocatalysis.

PubMed

Rizzo, Luigi

2009-06-15

In this study the potential application of TiO(2) photocatalysis as primary disinfection system of drinking water was investigated in terms of coliform bacteria inactivation and injury. As model water the effluent of biological denitrification unit for nitrate removal from groundwater, which is characterized by high organic matter and bacteria release, was used. The injury of photocatalysis on coliform bacteria was characterized by means of selective (mEndo) and less selective (mT7) culture media. Different catalyst loadings as well as photolysis and adsorption effects were investigated. Photocatalysis was effective in coliform bacteria inactivation (91-99% after 60 min irradiation time, depending on both catalyst loading and initial density of coliform bacteria detected by mEndo), although no total removal was observed after 60 min irradiation time. The contribution of adsorption mechanism was significant (60-98% after 60 min, depending on catalyst loading) compared to previous investigations probably due to the nature of source water rich in particulate organic matter and biofilm. Photocatalysis process did not result in any irreversible injury (98.8% being the higher injury) under investigated conditions, thus a bacteria regrowth may take place under optimum environment conditions if any final disinfection process (e.g., chlorine or chlorine dioxide) is not used.

Lipidomics as an important key for the identification of beer-spoilage bacteria.

PubMed

Řezanka, T; Matoulková, D; Benada, O; Sigler, K

2015-06-01

Electrospray ionization-tandem mass spectrometry (ESI-MS/MS) was used for characterizing intact plasmalogen phospholipid molecules in beer-spoilage bacteria. Identification of intact plasmalogens was carried out using collision-induced dissociation and the presence of suitable marker molecular species, both qualitative and quantitative, was determined in samples containing the anaerobic bacteria Megasphaera and Pectinatus. Using selected ion monitoring (SIM), this method had a limit of detection at 1 pg for the standard, i.e. 1-(1Z-octadecenyl)-2-oleoyl-sn-glycero-3-phosphoethanolamine and be linear in the range of four orders of magnitude from 2 pg to 20 ng. This technique was applied to intact plasmalogen extracts from the samples of contaminated and uncontaminated beer without derivatization and resulted in the identification of contamination of beer by Megasphaera and Pectinatus bacteria. The limit of detection was about 830 cells of anaerobic bacteria, i.e. bacteria containing natural cyclopropane plasmalogenes (c-p-19:0/15:0), which is the majority plasmalogen located in both Megasphaera and Pectinatus. The SIM ESI-MS method has been shown to be useful for the analysis of low concentration of plasmalogens in all biological samples, which were contaminated with anaerobic bacteria, e.g. juice, not only in beer. Significance and impact of the study: Electrospray ionization-tandem mass spectrometry (ESI-MS/MS) using collision-induced dissociation was used to characterize intact plasmalogen phospholipid molecules in beer-spoilage anaerobic bacteria Megasphaera and Pectinatus. Using selected ion monitoring (SIM), this method has a detection limit of 1 pg for the standard 1-(1Z-octadecenyl)-2-oleoyl-sn-glycero-3-phosphoethanolamine and is linear within four orders of magnitude (2 pg to 20 ng). The limit of detection was about 830 cells of bacteria containing natural cyclopropane plasmalogen (c-p-19:0/15:0). SIM ESI-MS method is useful for analyzing low concentrations of plasmalogens in biological samples contaminated with anaerobic bacteria, e.g. beer or juice. © 2015 The Society for Applied Microbiology.

Protein-linked glycans in periodontal bacteria: prevalence and role at the immune interface.

PubMed

Settem, Rajendra P; Honma, Kiyonobu; Stafford, Graham P; Sharma, Ashu

2013-10-17

Protein modification with complex glycans is increasingly being recognized in many pathogenic and non-pathogenic bacteria, and is now thought to be central to the successful life-style of those species in their respective hosts. This review aims to convey current knowledge on the extent of protein glycosylation in periodontal pathogenic bacteria and its role in the modulation of the host immune responses. The available data show that surface glycans of periodontal bacteria orchestrate dendritic cell cytokine responses to drive T cell immunity in ways that facilitate bacterial persistence in the host and induce periodontal inflammation. In addition, surface glycans may help certain periodontal bacteria protect against serum complement attack or help them escape immune detection through glycomimicry. In this review we will focus mainly on the generalized surface-layer protein glycosylation system of the periodontal pathogen Tannerella forsythia in shaping innate and adaptive host immunity in the context of periodontal disease. In addition, we will also review the current state of knowledge of surface protein glycosylation and its potential for immune modulation in other periodontal pathogens.

Immune system responses and fitness costs associated with consumption of bacteria in larvae of Trichoplusia ni

PubMed Central

Freitak, Dalial; Wheat, Christopher W; Heckel, David G; Vogel, Heiko

2007-01-01

Background Insects helped pioneer, and persist as model organisms for, the study of specific aspects of immunity. Although they lack an adaptive immune system, insects possess an innate immune system that recognizes and destroys intruding microorganisms. Its operation under natural conditions has not been well studied, as most studies have introduced microbes to laboratory-reared insects via artificial mechanical wounding. One of the most common routes of natural exposure and infection, however, is via food; thus, the role of dietary microbial communities in herbivorous insect immune system evolution invites study. Here, we examine the immune system response and consequences of exposing a lepidopteran agricultural pest to non-infectious microorganisms via simple oral consumption. Results Immune system response was compared between Trichoplusia ni larvae reared on diets with or without non-pathogenic bacteria (Escherichia coli and Micrococcus luteus). Two major immune response-related enzymatic activities responded to diets differently – phenoloxidase activity was inhibited in the bacteria-fed larvae, whereas general antibacterial activity was enhanced. Eight proteins were highly expressed in the hemolymph of the bacteria fed larvae, among them immune response related proteins arylphorin, apolipophorin III and gloverin. Expression response among 25 putative immune response-related genes were assayed via RT-qPCR. Seven showed more than fivefold up regulation in the presence of bacterial diet, with 22 in total being differentially expressed, among them apolipophorin III, cecropin, gallerimycin, gloverin, lysozyme, and phenoloxidase inhibiting enzyme. Finally, potential life-history trade-offs were studied, with pupation time and pupal mass being negatively affected in bacteria fed larvae. Conclusion The presence of bacteria in food, even if non-pathogenic, can trigger an immune response cascade with life history tradeoffs. Trichoplusia ni larvae are able to detect and respond to environmental microbes encountered in the diet, possibly even using midgut epithelial tissue as a sensing organ. Potential benefits of this immune system priming may outweigh the observed tradeoffs, as priming based on environmentally sensed bacterial may decrease risk of serious infection. These results show that food plant microbial communities represent a dynamic and unstudied part of the coevolutionary interactions between plants and their insect herbivores. PMID:18154650

Detection of γ-radiation and heavy metals using electrochemical bacterial-based sensor

NASA Astrophysics Data System (ADS)

Al-Shanawa, M.; Nabok, A.; Hashim, A.; Smith, T.; Forder, S.

2013-06-01

The main aim of this work is to develop a simple electrochemical sensor for detection of γ-radiation and heavy metals using bacteria. A series of DC and AC electrical measurements were carried out on samples of two types of bacteria, namely Escherichia coli and Deinococcus radiodurans. As a first step, a correlation between DC and AC electrical conductivity and bacteria concentration in solution was established. The study of the effect of γ-radiation and heavy metal ions (Cd2+) on DC and AC electrical characteristics of bacteria revealed a possibility of pattern recognition of the above inhibition factors.

Detection of Salmonella by indicator agar media and PCR as affected by alfalfa seed homogenates and native bacteria.

PubMed

Liao, C-H; Shollenberger, L M

2003-01-01

To investigate and prevent the undesirable effect of native bacteria and alfalfa seed homogenates on detection of Salmonella in alfalfa seeds by indicator agar media and polymerase chain reaction (PCR). The relative sensitivity of five indicator agar media, including modified semisolid RV (MSRV), xylose-lysine-Tergitol 4 (XLT4), Hektoen enteric agar (HEA), brilliant green agar (BGA) and bismuth sulphite agar (BSA), for detection of Salmonella in the presence of a large number of native bacteria from alfalfa seeds was examined. The detection limit as measured by the ratio between the numbers of native bacteria and Salmonella was estimated to be 10(6) to 1 for MSRV and 10(3) to 1 for XLT4, HEA, BGA or BSA. Presence of alfalfa seed homogenates markedly reduced the sensitivity of Salmonella detection by PCR. The minimal number of Salmonella detectable by PCR was determined to be 1-10 and 100-1000 CFU in the absence and presence of seed homogenate, respectively. Application of anti-Salmonella immunomagnetic beads permitted detection of 2-5 CFU of heat-injured cells in 25 g of seeds within 24 h by PCR. The MSRV medium is more sensitive than other indicator agars for detecting a small number of motile Salmonella in samples containing a large number of native bacteria. Application of immunomagnetic beads eliminates the PCR-inhibitory activity of seed homogenates and improves the detection of Salmonella in inoculated seeds. The results generated from this study will aid the seed distributors, sprout growers and public health officials to identify and recall the Salmonella-contaminated seed lots to be used for sprout production.

Adaptation of Tri-molecular fluorescence complementation allows assaying of regulatory Csr RNA-protein interactions in bacteria.

PubMed

Gelderman, Grant; Sivakumar, Anusha; Lipp, Sarah; Contreras, Lydia

2015-02-01

sRNAs play a significant role in controlling and regulating cellular metabolism. One of the more interesting aspects of certain sRNAs is their ability to make global changes in the cell by interacting with regulatory proteins. In this work, we demonstrate the use of an in vivo Tri-molecular Fluorescence Complementation assay to detect and visualize the central regulatory sRNA-protein interaction of the Carbon Storage Regulatory system in E. coli. The Carbon Storage Regulator consists primarily of an RNA binding protein, CsrA, that alters the activity of mRNA targets and of an sRNA, CsrB, that modulates the activity of CsrA. We describe the construction of a fluorescence complementation system that detects the interactions between CsrB and CsrA. Additionally, we demonstrate that the intensity of the fluorescence of this system is able to detect changes in the affinity of the CsrB-CsrA interaction, as caused by mutations in the protein sequence of CsrA. While previous methods have adopted this technique to study mRNA or RNA localization, this is the first attempt to use this technique to study the sRNA-protein interaction directly in bacteria. This method presents a potentially powerful tool to study complex bacterial RNA protein interactions in vivo. © 2014 Wiley Periodicals, Inc.

Molecular identification of coliform bacteria isolated from drinking water reservoirs with traditional methods and the Colilert-18 system.

PubMed

Kämpfer, Peter; Nienhüser, Anita; Packroff, Gabriele; Wernicke, Frank; Mehling, Arnd; Nixdorf, Katja; Fiedler, Stefanie; Kolauch, Claudia; Esser, Michael

2008-07-01

The accuracy of a traditional method (lactose utilization with acid and gas production) for the detection of coliform bacteria and E. coli was tested in comparison with method ISO 9308-1 (based on acid formation from lactose) and the Colilert-18 system (detection of beta-galactosidase). A total of 345 isolates were identified after isolation from water samples using API 20E strips. The Colilert-18 led to the highest number of positive findings (95% of the isolates were assigned to coliforms), whereas the ISO-9308-1 method resulted only in 29% coliform findings. With the traditional method only 15% were rated positive. Most of the isolates were identified by the API 20E system as Enterobacter spp. (species of the Enterobacter cloacae complex), Serratia spp., Citrobacter spp.and Klebsiella spp.; but species identification remained vague in several cases. A more detailed identification of 126 pure cultures by using 16S rRNA gene sequence analysis and analysis of the hsp60 gene resulted in the identification of Enterobacter nimipressuralis, E. amnigenus, E. asburiae, E. hormaechei, and Serratia fonticola as predominat coliforms. These species are beta-galactosidase positive, but show acid formation from lactose often after a prolonged incubation time. They are often not of fecal origin and may interfere with the ability to accurately detect coliforms of fecal origin.

Pathogen inactivation efficacy of Mirasol PRT System and Intercept Blood System for non-leucoreduced platelet-rich plasma-derived platelets suspended in plasma.

PubMed

Kwon, S Y; Kim, I S; Bae, J E; Kang, J W; Cho, Y J; Cho, N S; Lee, S W

2014-10-01

This study was conducted to evaluate the efficacy of pathogen inactivation (PI) in non-leucoreduced platelet-rich plasma-derived platelets suspended in plasma using the Mirasol PRT System and the Intercept Blood System. Platelets were pooled using the Acrodose PL system and separated into two aliquots for Mirasol and Intercept treatment. Four replicates of each viral strain were used for the evaluation. For bacteria, both low-titre (45-152 CFU/unit) inoculation and high-titre (7·34-10·18 log CFU/unit) inoculation with two replicates for each bacterial strain were used. Platelets with non-detectable bacterial growth and platelets inoculated with a low titre were stored for 5 days, and culture was performed with the BacT/ALERT system. The inactivation efficacy expressed as log reduction for Mirasol and Intercept systems for viruses was as follows: human immunodeficiency virus 1, ≥4·19 vs. ≥4·23; bovine viral diarrhoea virus, 1·83 vs. ≥6·03; pseudorabies virus, 2·73 vs. ≥5·20; hepatitis A virus, 0·62 vs. 0·76; and porcine parvovirus, 0·28 vs. 0·38. The inactivation efficacy for bacteria was as follows: Escherichia coli, 5·45 vs. ≥9·22; Staphylococcus aureus, 4·26 vs. ≥10·11; and Bacillus subtilis, 5·09 vs. ≥7·74. Postinactivation bacterial growth in platelets inoculated with a low titre of S. aureus or B. subtilis was detected only with Mirasol. Pathogen inactivation efficacy of Intercept for enveloped viruses was found to be satisfactory. Mirasol showed satisfactory inactivation efficacy for HIV-1 only. The two selected non-enveloped viruses were not inactivated by both systems. Inactivation efficacy of Intercept was more robust for all bacteria tested at high or low titres. © 2014 International Society of Blood Transfusion.

Substantial Expansion of Detectable Size Range in Ionic Current Sensing through Pores by Using a Microfluidic Bridge Circuit.

PubMed

Yasaki, Hirotoshi; Yasui, Takao; Yanagida, Takeshi; Kaji, Noritada; Kanai, Masaki; Nagashima, Kazuki; Kawai, Tomoji; Baba, Yoshinobu

2017-10-11

Measuring ionic currents passing through nano- or micropores has shown great promise for the electrical discrimination of various biomolecules, cells, bacteria, and viruses. However, conventional measurements have shown there is an inherent limitation to the detectable particle volume (1% of the pore volume), which critically hinders applications to real mixtures of biomolecule samples with a wide size range of suspended particles. Here we propose a rational methodology that can detect samples with the detectable particle volume of 0.01% of the pore volume by measuring a transient current generated from the potential differences in a microfluidic bridge circuit. Our method substantially suppresses the background ionic current from the μA level to the pA level, which essentially lowers the detectable particle volume limit even for relatively large pore structures. Indeed, utilizing a microscale long pore structure (volume of 5.6 × 10 4 aL; height and width of 2.0 × 2.0 μm; length of 14 μm), we successfully detected various samples including polystyrene nanoparticles (volume: 4 aL), bacteria, cancer cells, and DNA molecules. Our method will expand the applicability of ionic current sensing systems for various mixed biomolecule samples with a wide size range, which have been difficult to measure by previously existing pore technologies.

Protein Chips for Detection of Salmonella spp. from Enrichment Culture

PubMed Central

Poltronieri, Palmiro; Cimaglia, Fabio; De Lorenzis, Enrico; Chiesa, Maurizio; Mezzolla, Valeria; Reca, Ida Barbara

2016-01-01

Food pathogens are the cause of foodborne epidemics, therefore there is a need to detect the pathogens in food productions rapidly. A pre-enrichment culture followed by selective agar plating are standard detection methods. Molecular methods such as qPCR have provided a first rapid protocol for detection of pathogens within 24 h of enrichment culture. Biosensors also may provide a rapid tool to individuate a source of Salmonella contamination at early times of pre-enrichment culture. Forty mL of Salmonella spp. enrichment culture were processed by immunoseparation using the Pathatrix, as in AFNOR validated qPCR protocols. The Salmonella biosensor combined with immunoseparation showed a limit of detection of 100 bacteria/40 mL, with a 400 fold increase to previous results. qPCR analysis requires processing of bead-bound bacteria with lysis buffer and DNA clean up, with a limit of detection of 2 cfu/50 μL. Finally, a protein chip was developed and tested in screening and identification of 5 common pathogen species, Salmonella spp., E. coli, S. aureus, Campylobacter spp. and Listeria spp. The protein chip, with high specificity in species identification, is proposed to be integrated into a Lab-on-Chip system, for rapid and reproducible screening of Salmonella spp. and other pathogen species contaminating food productions. PMID:27110786

Linked Hydrologic-Hydrodynamic Model Framework to Forecast Impacts of Rivers on Beach Water Quality

NASA Astrophysics Data System (ADS)

Anderson, E. J.; Fry, L. M.; Kramer, E.; Ritzenthaler, A.

2014-12-01

The goal of NOAA's beach quality forecasting program is to use a multi-faceted approach to aid in detection and prediction of bacteria in recreational waters. In particular, our focus has been on the connection between tributary loads and bacteria concentrations at nearby beaches. While there is a clear link between stormwater runoff and beach water quality, quantifying the contribution of river loadings to nearshore bacterial concentrations is complicated due to multiple processes that drive bacterial concentrations in rivers as well as those processes affecting the fate and transport of bacteria upon exiting the rivers. In order to forecast potential impacts of rivers on beach water quality, we developed a linked hydrologic-hydrodynamic water quality framework that simulates accumulation and washoff of bacteria from the landscape, and then predicts the fate and transport of washed off bacteria from the watershed to the coastal zone. The framework includes a watershed model (IHACRES) to predict fecal indicator bacteria (FIB) loadings to the coastal environment (accumulation, wash-off, die-off) as a function of effective rainfall. These loadings are input into a coastal hydrodynamic model (FVCOM), including a bacteria transport model (Lagrangian particle), to simulate 3D bacteria transport within the coastal environment. This modeling system provides predictive tools to assist local managers in decision-making to reduce human health threats.

Detection of Mannitol Formation by Bacteria

PubMed Central

Chalfan, Y.; Levy, R.; Mateles, R. I.

1975-01-01

A test is described by means of which formation of mannitol from fructose by lactic acid bacteria can be readily detected. The test is based on removal of interference of residual fructose by dehydration with hydrochloric acid followed by thin-layer chromatography. PMID:1101827

Rapid fluorescence detection of pathogenic bacteria using magnetic enrichment technique combined with magnetophoretic chromatography.

PubMed

Che, Yulan; Xu, Yi; Wang, Renjie; Chen, Li

2017-08-01

A rapid and sensitive analytical method was developed to detect pathogenic bacteria which combined magnetic enrichment, fluorescence labeling with polyethylene glycol (PEG) magnetophoretic chromatography. As pathogenic bacteria usually exist in complex matrixes at low concentration, an efficient enrichment is essential for diagnosis. In order to capture series types of pathogenic bacteria in samples, amino-modified magnetic nanoparticles (Fe 3 O 4 @SiO 2 -NH 2 ) were prepared for efficient enrichment by the electrostatic interaction with pathogenic bacteria. It was shown that the capture efficiency reached up to 95.4% for Escherichia coli (E. coli). Furthermore, quantitative analysis of the bacteria was achieved by using acridine orange (AO) as a fluorescence probe for the captured E. coli due to its ability of staining series types of bacteria and rapid labeling. In order to remove the free magnetic nanoparticles and redundant fluorescent reagent, the labeled suspension was poured into a PEG separation column and was separated by applying an external magnetic field. The presence of 100 cfu mL -1 E. coli could be detected for semi-quantitative analysis by observing the separation column with the naked eye, and the concentration could be further evaluated by fluorescence detection. All the above processes were finished within 80 min. It was demonstrated that a good linear relationship existed between the fluorescence intensity and the concentration of E. coli ranging from 10 2 to 10 6  cfu mL -1 , with a detection limit of 100 cfu mL -1 when E. coli acted as target bacteria. The recovery rate of E. coli was 93.6∼102.0% in tap water and cooked meat samples, and the RSD was lower than 7% (n = 6); the result coincided with the conventional plate count method. Graphical abstract ᅟ.

Rapid direct methods for enumeration of specific, active bacteria in water and biofilms

NASA Technical Reports Server (NTRS)

McFeters, G. A.; Pyle, B. H.; Lisle, J. T.; Broadaway, S. C.

1999-01-01

Conventional methods for detecting indicator and pathogenic bacteria in water may underestimate the actual population due to sublethal environmental injury, inability of the target bacteria to take up nutrients and other physiological factors which reduce bacterial culturability. Rapid and direct methods are needed to more accurately detect and enumerate active bacteria. Such a methodological advance would provide greater sensitivity in assessing the microbiological safety of water and food. The principle goal of this presentation is to describe novel approaches we have formulated for the rapid and simultaneous detection of bacteria plus the determination of their physiological activity in water and other environmental samples. The present version of our method involves the concentration of organisms by membrane filtration or immunomagnetic separation and combines an intracellular fluorochrome (CTC) for assessment of respiratory activity plus fluorescent-labelled antibody detection of specific bacteria. This approach has also been successfully used to demonstrate spatial and temporal heterogeneities of physiological activities in biofilms when coupled with cryosectioning. Candidate physiological stains include those capable of determining respiratory activity, membrane potential, membrane integrity, growth rate and cellular enzymatic activities. Results obtained thus far indicate that immunomagnetic separation can provide a high degree of sensitivity in the recovery of seeded target bacteria (Escherichia coli O157:H7) in water and hamburger. The captured and stained target bacteria are then enumerated by either conventional fluorescence microscopy or ChemScan(R), a new instrument that is very sensitive and rapid. The ChemScan(R) laser scanning instrument (Chemunex, Paris, France) provides the detection of individual fluorescently labelled bacterial cells using three emission channels in less than 5 min. A high degree of correlation has been demonstrated between results obtained with the ChemScan and traditional plate counts of mixed natural bacterial populations in water. The continuing evolution of these methods will be valuable in the rapid and accurate analysis of environmental samples.

Surface-enhanced Raman spectroscopy introduced into the International Standard Organization (ISO) regulations as an alternative method for detection and identification of pathogens in the food industry.

PubMed

Witkowska, Evelin; Korsak, Dorota; Kowalska, Aneta; Księżopolska-Gocalska, Monika; Niedziółka-Jönsson, Joanna; Roźniecka, Ewa; Michałowicz, Weronika; Albrycht, Paweł; Podrażka, Marta; Hołyst, Robert; Waluk, Jacek; Kamińska, Agnieszka

2017-02-01

We show that surface-enhanced Raman spectroscopy (SERS) coupled with principal component analysis (PCA) can serve as a fast, reliable, and easy method for detection and identification of food-borne bacteria, namely Salmonella spp., Listeria monocytogenes, and Cronobacter spp., in different types of food matrices (salmon, eggs, powdered infant formula milk, mixed herbs, respectively). The main aim of this work was to introduce the SERS technique into three ISO (6579:2002; 11290-1:1996/A1:2004; 22964:2006) standard procedures required for detection of these bacteria in food. Our study demonstrates that the SERS technique is effective in distinguishing very closely related bacteria within a genus grown on solid and liquid media. The advantages of the proposed ISO-SERS method for bacteria identification include simplicity and reduced time of analysis, from almost 144 h required by standard methods to 48 h for the SERS-based approach. Additionally, PCA allows one to perform statistical classification of studied bacteria and to identify the spectrum of an unknown sample. Calculated first and second principal components (PC-1, PC-2) account for 96, 98, and 90% of total variance in the spectra and enable one to identify the Salmonella spp., L. monocytogenes, and Cronobacter spp., respectively. Moreover, the presented study demonstrates the excellent possibility for simultaneous detection of analyzed food-borne bacteria in one sample test (98% of PC-1 and PC-2) with a goal of splitting the data set into three separated clusters corresponding to the three studied bacteria species. The studies described in this paper suggest that SERS represents an alternative to standard microorganism diagnostic procedures. Graphical Abstract New approach of the SERS strategy for detection and identification of food-borne bacteria, namely S. enterica, L. monocytogenes, and C. sakazakii in selected food matrices.

Centrifugal sedimentation immunoassays for multiplexed detection of enteric bacteria in ground water

PubMed Central

Litvinov, Julia; Moen, Scott T.; Koh, Chung-Yan; Singh, Anup K.

2016-01-01

Waterborne pathogens pose significant threat to the global population and early detection plays an important role both in making drinking water safe, as well as in diagnostics and treatment of water-borne diseases. We present an innovative centrifugal sedimentation immunoassay platform for detection of bacterial pathogens in water. Our approach is based on binding of pathogens to antibody-functionalized capture particles followed by sedimentation of the particles through a density-media in a microfluidic disk. Beads at the distal end of the disk are imaged to quantify the fluorescence and determine the bacterial concentration. Our platform is fast (20 min), can detect as few as ∼10 bacteria with minimal sample preparation, and can detect multiple pathogens simultaneously. The platform was used to detect a panel of enteric bacteria (Escherichia coli, Salmonella typhimurium, Shigella, Listeria, and Campylobacter) spiked in tap and ground water samples. PMID:26858815

Inferring Models of Bacterial Dynamics toward Point Sources

PubMed Central

Jashnsaz, Hossein; Nguyen, Tyler; Petrache, Horia I.; Pressé, Steve

2015-01-01

Experiments have shown that bacteria can be sensitive to small variations in chemoattractant (CA) concentrations. Motivated by these findings, our focus here is on a regime rarely studied in experiments: bacteria tracking point CA sources (such as food patches or even prey). In tracking point sources, the CA detected by bacteria may show very large spatiotemporal fluctuations which vary with distance from the source. We present a general statistical model to describe how bacteria locate point sources of food on the basis of stochastic event detection, rather than CA gradient information. We show how all model parameters can be directly inferred from single cell tracking data even in the limit of high detection noise. Once parameterized, our model recapitulates bacterial behavior around point sources such as the “volcano effect”. In addition, while the search by bacteria for point sources such as prey may appear random, our model identifies key statistical signatures of a targeted search for a point source given any arbitrary source configuration. PMID:26466373

Imaging early pathogenesis of bubonic plague: are neutrophils commandeered for lymphatic transport of bacteria?

PubMed

Bland, David M; Anderson, Deborah M

2013-11-05

Vector-borne infections begin in the dermis when a pathogen is introduced by an arthropod during a blood meal. Several barriers separate an invading pathogen from its replicative niche, including phagocytic cells in the dermis that activate immunity by engulfing would-be pathogens and migrating to the lymph node. In addition, neutrophils circulating in the blood are rapidly recruited when the dermal barriers are penetrated. For flea-borne disease, no insect-encoded immune-suppressive molecules have yet been described that might influence the establishment of infection, leaving the bacteria on their own to defend against the mammalian immune system. Shortly after a flea transmits Yersinia pestis to a mammalian host, the bacteria are transported to the lymph node, where they grow logarithmically and later spread systemically. Even a single cell of Y. pestis can initiate a lethal case of plague. In their article, J. G. Shannon et al. [mBio 4(5):e00170-13, 2013, doi:10.1128/mBio.00170-13] used intravital microscopy to visualize trafficking of Y. pestis in transgenic mice in vivo, which allowed them to examine interactions between bacteria and specific immune cells. Bacteria appeared to preferentially interact with neutrophils but had no detectable interactions with dendritic cells. These findings suggest that Y. pestis infection of neutrophils not only prevents their activation but may even result in their return to circulation and migration to distal sites.

Autoimmunity: a decision theory model.

PubMed Central

Morris, J A

1987-01-01

Concepts from statistical decision theory were used to analyse the detection problem faced by the body's immune system in mounting immune responses to bacteria of the normal body flora. Given that these bacteria are potentially harmful, that there can be extensive cross reaction between bacterial antigens and host tissues, and that the decisions are made in uncertainty, there is a finite chance of error in immune response leading to autoimmune disease. A model of ageing in the immune system is proposed that is based on random decay in components of the decision process, leading to a steep age dependent increase in the probability of error. The age incidence of those autoimmune diseases which peak in early and middle life can be explained as the resultant of two processes: an exponentially falling curve of incidence of first contact with common bacteria, and a rapidly rising error function. Epidemiological data on the variation of incidence with social class, sibship order, climate and culture can be used to predict the likely site of carriage and mode of spread of the causative bacteria. Furthermore, those autoimmune diseases precipitated by common viral respiratory tract infections might represent reactions to nasopharyngeal bacterial overgrowth, and this theory can be tested using monoclonal antibodies to search the bacterial isolates for cross reacting antigens. If this model is correct then prevention of autoimmune disease by early exposure to low doses of bacteria might be possible. PMID:3818985

[Anaerobic biodegradation of phthalic acid esters (Paes) in municipal sludge].

PubMed

Liang, Zhi-Feng; Zhou, Wen; Lin, Qing-Qi; Yang, Xiu-Hong; Wang, Shi-Zhong; Cai, Xin-De; Qiu, Rong-Liang

2014-04-01

Phthalic acid esters (PAEs), a class of organic pollutants with potent endocrine-disrupting properties, are widely present in municipal sludge. Study of PAEs biodegradation under different anaerobic biological treatment processes of sludge is, therefore, essential for a safe use of sludge in agricultural practice. In this study, we selected two major sludge PAEs, i.e. di-n-butyl phthalate (DBP) and di-(2-enthylhexyl) phthalate (DEHP), to investigate their biodegradation behaviors in an anaerobic sludge digestion system and a fermentative hydrogen production system. The possible factors influencing PAEs biodegradation in relation to changes of sludge properties were also discussed. The results showed that the biodegradation of DBP reached 99.6% within 6 days, while that of DEHP was 46.1% during a 14-day incubation period in the anaerobic digestion system. By comparison, only 19.5% of DBP was degraded within 14 days in the fermentative hydrogen production system, while no degradation was detected for DEHP. The strong inhibition of the degradation of both PAEs in the fermentative hydrogen production system was ascribed to the decreases in microbial biomass and ratios of gram-positive bacteria/gram-negative bacteria and fungi/ bacteria, and the increase of concentrations of volatile fatty acids (e. g. acetic acid, propionic acid and butyric acid) during the fermentative hydrogen-producing process.

Algicidal and growth-inhibiting bacteria associated with seagrass and macroalgae beds in Puget Sound, WA, USA.

PubMed

Inaba, Nobuharu; Trainer, Vera L; Onishi, Yuka; Ishii, Ken-Ichiro; Wyllie-Echeverria, Sandy; Imai, Ichiro

2017-02-01

The algicidal and growth-inhibiting bacteria associated with seagrasses and macroalgae were characterized during the summer of 2012 and 2013 throughout Puget Sound, WA, USA. In 2012, Heterosigma akashiwo-killing bacteria were observed in concentrations of 2.8×10 6 CFUg -1 wet in the outer organic layer (biofilm) on the common eelgrass (Zostera marina) in north Padilla Bay. Bacteria that inhibited the growth of Alexandrium tamarense were detected within the biofilm formed on the eelgrass canopy at Dumas Bay and North Bay at densities of ∼10 8 CFUg -1 wet weight. Additionally, up to 4100CFUmL -1 of algicidal and growth-inhibiting bacteria affecting both A. tamarense and H. akashiwo were detected in seawater adjacent to seven different eelgrass beds. In 2013, H. akashiwo-killing bacteria were found on Z. marina and Ulva lactuca with the highest densities of ∼10 8 CFUg -1 wet weight at Shallow Bay, Sucia Island. Bacteria that inhibited the growth of H. akashiwo and A. tamarense were also detected on Z. marina and Z. japonica at central Padilla Bay. Heterosigma akashiwo cysts were detected at a concentration of 3400cystsg -1 wet weight in the sediment from Westcott Bay (northern San Juan Island), a location where eelgrass disappeared in 2002. These findings provide new insights on the ecology of algicidal and growth-inhibiting bacteria, and suggest that seagrass and macroalgae provide an environment that may influence the abundance of harmful algae in this region. This work highlights the importance of protection and restoration of native seagrasses and macroalgae in nearshore environments, in particular those regions where shellfish restoration initiatives are in place to satisfy a growing demand for seafood. Copyright © 2016 Elsevier B.V. All rights reserved.

Phage lytic enzymes: a history.

PubMed

Trudil, David

2015-02-01

There are many recent studies regarding the efficacy of bacteriophage-related lytic enzymes: the enzymes of 'bacteria-eaters' or viruses that infect bacteria. By degrading the cell wall of the targeted bacteria, these lytic enzymes have been shown to efficiently lyse Gram-positive bacteria without affecting normal flora and non-related bacteria. Recent studies have suggested approaches for lysing Gram-negative bacteria as well (Briersa Y, et al., 2014). These enzymes include: phage-lysozyme, endolysin, lysozyme, lysin, phage lysin, phage lytic enzymes, phageassociated enzymes, enzybiotics, muralysin, muramidase, virolysin and designations such as Ply, PAE and others. Bacteriophages are viruses that kill bacteria, do not contribute to antimicrobial resistance, are easy to develop, inexpensive to manufacture and safe for humans, animals and the environment. The current focus on lytic enzymes has been on their use as anti-infectives in humans and more recently in agricultural research models. The initial translational application of lytic enzymes, however, was not associated with treating or preventing a specific disease but rather as an extraction method to be incorporated in a rapid bacterial detection assay (Bernstein D, 1997).The current review traces the translational history of phage lytic enzymes-from their initial discovery in 1986 for the rapid detection of group A streptococcus in clinical specimens to evolving applications in the detection and prevention of disease in humans and in agriculture.

Optical sensing: recognition elements and devices

NASA Astrophysics Data System (ADS)

Gauglitz, Guenter G.

2012-09-01

The requirements in chemical and biochemical sensing with respect to recognition elements, avoiding non-specific interactions, and high loading of the surface for detection of low concentrations as well as optimized detection systems are discussed. Among the many detection principles the optical techniques are classified. Methods using labeled compounds like Total Internal Reflection Fluorescence (TIRF) and direct optical methods like micro reflectometry or refractometry are discussed in comparison. Reflectometric Interference Spectroscopy (RIfS) is presented as a robust simple method for biosensing. As applications, trace analysis of endocrine disruptors in water, hormones in food, detection of viruses and bacteria in food and clinical diagnostics are discussed.

Microtiter plate-based antibody microarrays for bacteria and toxins

USDA-ARS?s Scientific Manuscript database

Research has focused on the development of rapid biosensor-based, high-throughput, and multiplexed detection of pathogenic bacteria in foods. Specifically, antibody microarrays in 96-well microtiter plates have been generated for the purpose of selective detection of Shiga toxin-producing E. coli (...

A METHOD TO DETECT VIABLE HELICOBACTER PYLORI BACTERIA IN GROUNDWATER

EPA Science Inventory

The inability to detect the presence of viable Helicobacter pylori bacteria in environmental waters has hindered the public health community in assessing the role water may playin the transmission of this pathogen. This work describes a cultural enrichment method coupled with an...

Detection of Spiroplasma citri by droplet digital PCR

USDA-ARS?s Scientific Manuscript database

Spiroplasmas are motile, helical bacteria belonging to the Class Mollicutes, a group of prokaryotics with no cell wall and phylogenetically related to Gram-positive bacteria. Spiroplasma citri is the first-cultured spiroplasma and causal agent of citrus stubborn disease (CSD). Detection of CSD is di...

Optimization of an enhanced ceramic micro-filter for concentrating E.coli in water

NASA Astrophysics Data System (ADS)

Zhang, Yushan; Guo, Tianyi; Xu, Changqing; Hong, Lingcheng

2017-02-01

Recently lower limit of detection (LOD) is necessary for rapid bacteria detection and analysis applications in clinical practices and daily life. A critical pre-conditioning step for these applications is bacterial concentration, especially for low level of pathogens. Sample volume can be largely reduced with an efficient pre-concentration process. Some approaches such as hollow-fiber ultra-filtration and electrokinetic technique have been applied to bacterial concentration. Since none of these methods can provide a concentrating method with a stable recovery efficiency, bacterial concentration still remains challenging Ceramic micro- filter can be used to concentrate the bacteria but the cross flow system keeps the bacteria in suspension. Similar harvesting bacteria using ultra-filtration showed an average recovery efficiency of 43% [1] and other studies achieved recovery rates greater than 50% [2]. In this study, an enhanced ceramic micro-filter with 0.14 μm pore size was proposed and demonstrated to optimize the concentration of E.coli. A high recovery rate (mean value >90%) and a high volumetric concentration ratio (>100) were achieved. Known quantities (104 to 106 CFU/ml) of E.coli cells were spiked to different amounts of phosphate buffered saline (0.1 to 1 L), and then concentrated to a final retentate of 5 ml to 10 ml. An average recovery efficiency of 95.3% with a standard deviation of 5.6% was achieved when the volumetric con- centration ratio was 10. No significant recovery rate loss was indicated when the volumetric concentration ratio reached up to 100. The effects of multiple parameters on E.coli recovery rate were also studied. The obtained results indicated that the optimized ceramic micro- filtration system can successfully concentrate E.coli cells in water with an average recovery rate of 90.8%.

Differential plating medium for quantitative detection of histamine-producing bacteria.

PubMed Central

Niven, C F; Jeffrey, M B; Corlett, D A

1981-01-01

A histidine-containing agar medium has been devised for quantitative detection of histamine-producing bacteria that are alleged to be associated with scombroid fish poisoning outbreaks. The responsible bacteria produce a marked pH change in the agar, with attendant color change of pH indicator adjacent to the colonies, thus facilitating their recognition. Proteus morganii and Klebsiella pneumoniae were the two most common histidine-decarboxylating species isolated from scombroid fish and mahi mahi. PMID:7013698

Application of immobilized synthetic anti-lipopolysaccharide peptides for the isolation and detection of bacteria.

PubMed

Sandetskaya, N; Engelmann, B; Brandenburg, K; Kuhlmeier, D

2015-08-01

The molecular detection of microorganisms in liquid samples generally requires their enrichment or isolation. The aim of our study was to evaluate the capture and pre-concentration of bacteria by immobilized particular cationic antimicrobial peptides, called synthetic anti-lipopolysaccharide peptides (SALP). For the proof-of-concept and screening of different SALP, the peptides were covalently immobilized on glass slides, and the binding of bacteria was confirmed by microscopic examination of the slides or their scanning, in case of fluorescent bacterial cells. The most efficient SALP was further tethered to magnetic beads. SALP beads were used for the magnetic capture of Escherichia coli in liquid samples. The efficiency of this strategy was evaluated using polymerase chain reaction (PCR). Covalently immobilized SALP were capable of capturing bacteria in liquid samples. However, PCR was hampered by the unspecific binding of DNA to the positively charged peptide. We developed a method for DNA recovery by the enzymatic digestion of the peptide, which allowed for a successful PCR, though the method had its own adverse impact on the detection and, thus, did not allow for the reliable quantitative analysis of the pathogen enrichment. Immobilized SALP can be used as capture molecules for bacteria in liquid samples and can be recommended for the design of the assays or decontamination of the fluids. For the accurate subsequent detection of bacteria, DNA-independent methods should be used.

Establishment of HPC(R2A) for regrowth control in non-chlorinated distribution systems.

PubMed

Uhl, Wolfgang; Schaule, Gabriela

2004-05-01

Drinking water distributed without disinfection and without regrowth problems for many years may show bacterial regrowth when the residence time and/or temperature in the distribution system increases or when substrate and/or bacterial concentration in the treated water increases. An example of a regrowth event in a major German city is discussed. Regrowth of HPC bacteria occurred unexpectedly at the end of a very hot summer. No pathogenic or potentially pathogenic bacteria were identified. Increased residence times in the distribution system and temperatures up to 25 degrees C were identified as most probable causes and the regrowth event was successfully overcome by changing flow regimes and decreasing residence times. Standard plate counts of HPC bacteria using the spread plate technique on nutrient rich agar according to German Drinking Water Regulations (GDWR) had proven to be a very good indicator of hygienically safe drinking water and to demonstrate the effectiveness of water treatment. However, the method proved insensitive for early regrowth detection. Regrowth experiments in the lab and sampling of the distribution system during two summers showed that spread plate counts on nutrient-poor R2A agar after 7-day incubation yielded 100 to 200 times higher counts. Counts on R2A after 3-day incubation were three times less than after 7 days. As the precision of plate count methods is very poor for counts less than 10 cfu/plate, a method yielding higher counts is better suited to detect upcoming regrowth than a method yielding low counts. It is shown that for the identification of regrowth events HPC(R2A) gives a further margin of about 2 weeks for reaction before HPC(GDWR). Copyright 2003 Elsevier B.V.

Nucleic Acid Immunity.

PubMed

Hartmann, G

2017-01-01

Organisms throughout biology need to maintain the integrity of their genome. From bacteria to vertebrates, life has established sophisticated mechanisms to detect and eliminate foreign genetic material or to restrict its function and replication. Tremendous progress has been made in the understanding of these mechanisms which keep foreign or unwanted nucleic acids from viruses or phages in check. Mechanisms reach from restriction-modification systems and CRISPR/Cas in bacteria and archaea to RNA interference and immune sensing of nucleic acids, altogether integral parts of a system which is now appreciated as nucleic acid immunity. With inherited receptors and acquired sequence information, nucleic acid immunity comprises innate and adaptive components. Effector functions include diverse nuclease systems, intrinsic activities to directly restrict the function of foreign nucleic acids (e.g., PKR, ADAR1, IFIT1), and extrinsic pathways to alert the immune system and to elicit cytotoxic immune responses. These effects act in concert to restrict viral replication and to eliminate virus-infected cells. The principles of nucleic acid immunity are highly relevant for human disease. Besides its essential contribution to antiviral defense and restriction of endogenous retroelements, dysregulation of nucleic acid immunity can also lead to erroneous detection and response to self nucleic acids then causing sterile inflammation and autoimmunity. Even mechanisms of nucleic acid immunity which are not established in vertebrates are relevant for human disease when they are present in pathogens such as bacteria, parasites, or helminths or in pathogen-transmitting organisms such as insects. This review aims to provide an overview of the diverse mechanisms of nucleic acid immunity which mostly have been looked at separately in the past and to integrate them under the framework nucleic acid immunity as a basic principle of life, the understanding of which has great potential to advance medicine. © 2017 Elsevier Inc. All rights reserved.

Drinking water biofilms on copper and stainless steel exhibit specific molecular responses towards different disinfection regimes at waterworks.

PubMed

Jungfer, Christina; Friedrich, Frank; Varela Villarreal, Jessica; Brändle, Katharina; Gross, Hans-Jürgen; Obst, Ursula; Schwartz, Thomas

2013-09-01

Biofilms growing on copper and stainless steel substrata in natural drinking water were investigated. A modular pilot-scale distribution facility was installed at four waterworks using different raw waters and disinfection regimes. Three-month-old biofilms were analysed using molecular biology and microscopy methods. High total cell numbers, low counts of actively respiring cells and low numbers of cultivable bacteria indicated the high abundance of viable but not cultivable bacteria in the biofilms. The expression of the recA SOS responsive gene was detected and underlined the presence of transcriptionally active bacteria within the biofilms. This effect was most evident after UV disinfection, UV oxidation and UV disinfection with increased turbidity at waterworks compared to chemically treated and non-disinfected systems. Furthermore, live/dead staining techniques and environmental scanning electron microscopy imaging revealed the presence of living and intact bacteria in biofilms on copper substrata. Cluster analyses of DGGE profiles demonstrated differences in the composition of biofilms on copper and steel materials.

Phytoplankton production systems in a shellfish hatchery: variations of the bacterial load and diversity of vibrios.

PubMed

Dubert, J; Fernández-Pardo, A; Nóvoa, S; Barja, J L; Prado, S

2015-06-01

Outbreaks of disease caused by some Vibrio species represent the main production bottleneck in shellfish hatcheries. Although the phytoplankton used as food is one of the main sources of bacteria, studies of the associated bacterial populations, specifically vibrios, are scarce. The aim of the study was the microbiological monitoring of the microalgae as the first step in assessing the risk disease for bivalve cultures. Two phytoplankton production systems were sampled weekly throughout 1-year period in a bivalve hatchery. Quantitative analysis revealed high levels of marine heterotrophic bacteria in both systems throughout the study. Presumptive vibrios were detected occasionally and at low concentrations. In most of the cases, they belonged to the Splendidus and Harveyi clades. The early detection of vibrios in the microalgae may be the key for a successful bivalve culture. Their abundance and diversity were affected by factors related to the hatchery environment. This work represents the first long study where the presence of vibrios was evaluated rigorously in phytoplankton production systems and provides a suitable microbiological protocol to control and guarantee the quality of the algal cultures to avoid the risk of transferring potential pathogens to shellfish larvae and/or broodstock. © 2015 The Society for Applied Microbiology.

Bioluminescence Resonance Energy Transfer System for Measuring Dynamic Protein-Protein Interactions in Bacteria

PubMed Central

Cui, Boyu; Wang, Yao; Song, Yunhong; Wang, Tietao; Li, Changfu; Wei, Yahong

2014-01-01

ABSTRACT Protein-protein interactions are important for virtually every biological process, and a number of elegant approaches have been designed to detect and evaluate such interactions. However, few of these methods allow the detection of dynamic and real-time protein-protein interactions in bacteria. Here we describe a bioluminescence resonance energy transfer (BRET) system based on the bacterial luciferase LuxAB. We found that enhanced yellow fluorescent protein (eYFP) accepts the emission from LuxAB and emits yellow fluorescence. Importantly, BRET occurred when LuxAB and eYFP were fused, respectively, to the interacting protein pair FlgM and FliA. Furthermore, we observed sirolimus (i.e., rapamycin)-inducible interactions between FRB and FKBP12 and a dose-dependent abolishment of such interactions by FK506, the ligand of FKBP12. Using this system, we showed that osmotic stress or low pH efficiently induced multimerization of the regulatory protein OmpR and that the multimerization induced by low pH can be reversed by a neutralizing agent, further indicating the usefulness of this system in the measurement of dynamic interactions. This method can be adapted to analyze dynamic protein-protein interactions and the importance of such interactions in bacterial processes such as development and pathogenicity. PMID:24846380

Bioluminescence resonance energy transfer system for measuring dynamic protein-protein interactions in bacteria.

PubMed

Cui, Boyu; Wang, Yao; Song, Yunhong; Wang, Tietao; Li, Changfu; Wei, Yahong; Luo, Zhao-Qing; Shen, Xihui

2014-05-20

Protein-protein interactions are important for virtually every biological process, and a number of elegant approaches have been designed to detect and evaluate such interactions. However, few of these methods allow the detection of dynamic and real-time protein-protein interactions in bacteria. Here we describe a bioluminescence resonance energy transfer (BRET) system based on the bacterial luciferase LuxAB. We found that enhanced yellow fluorescent protein (eYFP) accepts the emission from LuxAB and emits yellow fluorescence. Importantly, BRET occurred when LuxAB and eYFP were fused, respectively, to the interacting protein pair FlgM and FliA. Furthermore, we observed sirolimus (i.e., rapamycin)-inducible interactions between FRB and FKBP12 and a dose-dependent abolishment of such interactions by FK506, the ligand of FKBP12. Using this system, we showed that osmotic stress or low pH efficiently induced multimerization of the regulatory protein OmpR and that the multimerization induced by low pH can be reversed by a neutralizing agent, further indicating the usefulness of this system in the measurement of dynamic interactions. This method can be adapted to analyze dynamic protein-protein interactions and the importance of such interactions in bacterial processes such as development and pathogenicity. Real-time measurement of protein-protein interactions in prokaryotes is highly desirable for determining the roles of protein complex in the development or virulence of bacteria, but methods that allow such measurement are not available. Here we describe the development of a bioluminescence resonance energy transfer (BRET) technology that meets this need. The use of endogenous excitation light in this strategy circumvents the requirement for the sophisticated instrument demanded by standard fluorescence resonance energy transfer (FRET). Furthermore, because the LuxAB substrate decanal is membrane permeable, the assay can be performed without lysing the bacterial cells, thus allowing the detection of protein-protein interactions in live bacterial cells. This BRET system added another useful tool to address important questions in microbiological studies. Copyright © 2014 Cui et al.

Endosafe(R)-Portable Test System (PTS)

NASA Technical Reports Server (NTRS)

Maule, Jake; Wainwright, Norm; Burbank, Dan

2005-01-01

The Portable Test System (PTS) is a hand-held device for monitoring the presence of potentially hazardous bacteria in the environment. It uses an immunological method derived from the horseshoe crab (Limulus polyphemus) to detect bacterial cell membranes and other molecular components of a cell. Further modifications of the PTS will allow detection of individual hazardous species of bacteria. This study was a follow-up of previous PTS and other immunological tests performed on the KC-135 during 2002-2003 (Maule et al., 2003, J. Gravit. Physiol.) and in the underwater habitat Aquarius during NEEMO 5 (Maule et al., 2005, Appl. Environ. Microbiol in prep.). The experiments described here were part of a final testing phase prior to use of the PTS on the International Space Station (ISS), scheduled for launch on 12A.1 on February 9th 2006. The specific aspects of PTS operation studied were those involving a fluid component: pumping, mixing, incubations and pipetting into the instrument. The PTS uses a stepper motor to move fluid along small channels, which may be affected by reduced gravity.

Cell death in a harmful algal bloom causing species Alexandrium tamarense upon an algicidal bacterium induction.

PubMed

Zhang, Huajun; Lv, Jinglin; Peng, Yun; Zhang, Su; An, Xinli; Xu, Hong; Zhang, Jun; Tian, Yun; Zheng, Wei; Zheng, Tianling

2014-09-01

Harmful algal blooms occur throughout the world, destroying aquatic ecosystems and threatening human health. The culture supernatant of the marine algicidal bacteria DHQ25 was able to lysis dinoflagellate Alexandrium tamarense. Loss of photosynthetic pigments, accompanied by a decline in Photosystem II (PSII) photochemical efficiency (Fv/Fm), in A. tamarense was detected under bacterial supernatant stress. Transmission electron microscope analysis showed obvious morphological modifications of chloroplast dismantling as a part of the algicidal process. The PSII electron transport chain was seriously blocked, with its reaction center damaged. This damage was detected in a relative transcriptional level of psbA and psbD genes, which encode the D1 and D2 proteins in the PSII reaction center. And the block in the electron transport chain of PSII might generate excessive reactive oxygen species (ROS) which could destroy the membrane system and pigment synthesis and activated enzymic antioxidant systems including superoxide dismutase (SOD) and catalase (CAT). This study indicated that marine bacteria with indirect algicidal activity played an important role in the changes of photosynthetic process in a harmful algal bloom species.

Mechanisms affecting water quality in an intermittent piped water supply.

PubMed

Kumpel, Emily; Nelson, Kara L

2014-01-01

Drinking water distribution systems throughout the world supply water intermittently, leaving pipes without pressure between supply cycles. Understanding the multiple mechanisms that affect contamination in these intermittent water supplies (IWS) can be used to develop strategies to improve water quality. To study these effects, we tested water quality in an IWS system with infrequent and short water delivery periods in Hubli-Dharwad, India. We continuously measured pressure and physicochemical parameters and periodically collected grab samples to test for total coliform and E. coli throughout supply cycles at 11 sites. When the supply was first turned on, water with elevated turbidity and high concentrations of indicator bacteria was flushed out of pipes. At low pressures (<10 psi), elevated indicator bacteria were frequently detected even when there was a chlorine residual, suggesting persistent contamination had occurred through intrusion or backflow. At pressures between 10 and 17 psi, evidence of periodic contamination suggested that transient intrusion, backflow, release of particulates, or sloughing of biofilms from pipe walls had occurred. Few total coliform and no E. coli were detected when water was delivered with a chlorine residual and at pressures >17 psi.

Bench-to-bedside review: Quorum sensing and the role of cell-to-cell communication during invasive bacterial infection

PubMed Central

Asad, Shadaba; Opal, Steven M

2008-01-01

Bacteria communicate extensively with each other and employ a communal approach to facilitate survival in hostile environments. A hierarchy of cell-to-cell signaling pathways regulates bacterial growth, metabolism, biofilm formation, virulence expression, and a myriad of other essential functions in bacterial populations. The notion that bacteria can signal each other and coordinate their assault patterns against susceptible hosts is now well established. These signaling networks represent a previously unrecognized survival strategy by which bacterial pathogens evade antimicrobial defenses and overwhelm the host. These quorum sensing communication signals can transgress species barriers and even kingdom barriers. Quorum sensing molecules can regulate human transcriptional programs to the advantage of the pathogen. Human stress hormones and cytokines can be detected by bacterial quorum sensing systems. By this mechanism, the pathogen can detect the physiologically stressed host, providing an opportunity to invade when the patient is most vulnerable. These rather sophisticated, microbial communication systems may prove to be a liability to pathogens as they make convenient targets for therapeutic intervention in our continuing struggle to control microbial pathogens. PMID:19040778

Effects of various fertilization regimes on abundance and activity of anaerobic ammonium oxidation bacteria in rice-wheat cropping systems in China.

PubMed

Gu, Chao; Zhou, Huifang; Zhang, Qichun; Zhao, Yuhua; Di, Hongjie; Liang, Yongchao

2017-12-01

Anaerobic ammonium oxidation (anammox) is an important process in many marine and paddy ecosystems. However, few studies have reported on the contribution to the nitrogen cycle of anammox and its dynamics in rice-wheat cropping systems with different fertilization regimes. Here, isotope tracing and molecular techniques were used to determine the potential rates of anammox and their association with bacterial abundance, diversity, and activity. Rice-wheat cropping systems at two sites in Jiangsu Province, China were selected and the treatments at each site were: 1) no fertilization (CK), 2) 100% chemical fertilization (CF), 3) pig manure compost plus 50% chemical fertilization (PMCF), and 4) straw plus 100% chemical fertilization (SRCF). The results revealed that anammox bacteria with high abundance were detected in both the wheat and rice seasons. The abundance of anammox in PMCF treatment was higher than that in SRCF treatment in both Changshu and Jintan. Moreover, the abundance of anammox bacteria in CF treatment was significantly higher than that in CK in Changshu. Analysis of anammox hydrazine synthase β subunit (hzs-β) gene sequences showed that in the rice season, the anammox bacteria Ca. Brocadia, Ca. Scalindua, and Ca. Jettenia were present. In contrast, all of the anammox hydrazine oxidase (hzo) genes were affiliated with Ca. Brocadia, suggesting that hzs genes are more representative of anammox biological diversity compared to hzo. Sequences from the PMCF treatment where affiliated with both Ca. Jettenia and Ca. Brocadia, and showed the highest diversity. Anammox activity was detected in both the wheat and rice seasons, but there were significant differences between seasons. The anammox rates were in the range 0.34 to 1.04nmol dinitrogen gas∙g -1 dry soil∙h -1 , and 3.15 to 9.62% of dinitrogen gas emissions were attributed to anammox. However, no significant difference among the fertilizer treatments for anammox activity was found in the study. Copyright © 2017. Published by Elsevier B.V.

Protozoan Bacterivory and Escherichia coli Survival in Drinking Water Distribution Systems

PubMed Central

Sibille, I.; Sime-Ngando, T.; Mathieu, L.; Block, J. C.

1998-01-01

The development of bacterial communities in drinking water distribution systems leads to a food chain which supports the growth of macroorganisms incompatible with water quality requirements and esthetics. Nevertheless, very few studies have examined the microbial communities in drinking water distribution systems and their trophic relationships. This study was done to quantify the microbial communities (especially bacteria and protozoa) and obtain direct and indirect proof of protozoan feeding on bacteria in two distribution networks, one of GAC water (i.e., water filtered on granular activated carbon) and the other of nanofiltered water. The nanofiltered water-supplied network contained no organisms larger than bacteria, either in the water phase (on average, 5 × 107 bacterial cells liter−1) or in the biofilm (on average, 7 × 106 bacterial cells cm−2). No protozoa were detected in the whole nanofiltered water-supplied network (water plus biofilm). In contrast, the GAC water-supplied network contained bacteria (on average, 3 × 108 cells liter−1 in water and 4 × 107 cells cm−2 in biofilm) and protozoa (on average, 105 cells liter−1 in water and 103 cells cm−2 in biofilm). The water contained mostly flagellates (93%), ciliates (1.8%), thecamoebae (1.6%), and naked amoebae (1.1%). The biofilm had only ciliates (52%) and thecamoebae (48%). Only the ciliates at the solid-liquid interface of the GAC water-supplied network had a measurable grazing activity in laboratory test (estimated at 2 bacteria per ciliate per h). Protozoan ingestion of bacteria was indirectly shown by adding Escherichia coli to the experimental distribution systems. Unexpectedly, E. coli was lost from the GAC water-supplied network more rapidly than from the nanofiltered water-supplied network, perhaps because of the grazing activity of protozoa in GAC water but not in nanofiltered water. Thus, the GAC water-supplied network contained a functional ecosystem with well-established and structured microbial communities, while the nanofiltered water-supplied system did not. The presence of protozoa in drinking water distribution systems must not be neglected because these populations may regulate the autochthonous and allochthonous bacterial populations. PMID:9435076

Protozoan bacterivory and Escherichia coli survival in drinking water distribution systems.

PubMed

Sibille, I; Sime-Ngando, T; Mathieu, L; Block, J C

1998-01-01

The development of bacterial communities in drinking water distribution systems leads to a food chain which supports the growth of macroorganisms incompatible with water quality requirements and esthetics. Nevertheless, very few studies have examined the microbial communities in drinking water distribution systems and their trophic relationships. This study was done to quantify the microbial communities (especially bacteria and protozoa) and obtain direct and indirect proof of protozoan feeding on bacteria in two distribution networks, one of GAC water (i.e., water filtered on granular activated carbon) and the other of nanofiltered water. The nanofiltered water-supplied network contained no organisms larger than bacteria, either in the water phase (on average, 5 x 10(7) bacterial cells liter-1) or in the biofilm (on average, 7 x 10(6) bacterial cells cm-2). No protozoa were detected in the whole nanofiltered water-supplied network (water plus biofilm). In contrast, the GAC water-supplied network contained bacteria (on average, 3 x 10(8) cells liter-1 in water and 4 x 10(7) cells cm-2 in biofilm) and protozoa (on average, 10(5) cells liter-1 in water and 10(3) cells cm-2 in biofilm). The water contained mostly flagellates (93%), ciliates (1.8%), thecamoebae (1.6%), and naked amoebae (1.1%). The biofilm had only ciliates (52%) and thecamoebae (48%). Only the ciliates at the solid-liquid interface of the GAC water-supplied network had a measurable grazing activity in laboratory test (estimated at 2 bacteria per ciliate per h). Protozoan ingestion of bacteria was indirectly shown by adding Escherichia coli to the experimental distribution systems. Unexpectedly, E. coli was lost from the GAC water-supplied network more rapidly than from the nanofiltered water-supplied network, perhaps because of the grazing activity of protozoa in GAC water but not in nanofiltered water. Thus, the GAC water-supplied network contained a functional ecosystem with well-established and structured microbial communities, while the nanofiltered water-supplied system did not. The presence of protozoa in drinking water distribution systems must not be neglected because these populations may regulate the autochthonous and allochthonous bacterial populations.

Community Analysis of Biofilters Using Fluorescence In Situ Hybridization Including a New Probe for the Xanthomonas Branch of the Class Proteobacteria

PubMed Central

Friedrich, Udo; Naismith, Michèle M.; Altendorf, Karlheinz; Lipski, André

1999-01-01

Domain-, class-, and subclass-specific rRNA-targeted probes were applied to investigate the microbial communities of three industrial and three laboratory-scale biofilters. The set of probes also included a new probe (named XAN818) specific for the Xanthomonas branch of the class Proteobacteria; this probe is described in this study. The members of the Xanthomonas branch do not hybridize with previously developed rRNA-targeted oligonucleotide probes for the α-, β-, and γ-Proteobacteria. Bacteria of the Xanthomonas branch accounted for up to 4.5% of total direct counts obtained with 4′,6-diamidino-2-phenylindole. In biofilter samples, the relative abundance of these bacteria was similar to that of the γ-Proteobacteria. Actinobacteria (gram-positive bacteria with a high G+C DNA content) and α-Proteobacteria were the most dominant groups. Detection rates obtained with probe EUB338 varied between about 40 and 70%. For samples with high contents of gram-positive bacteria, these percentages were substantially improved when the calculations were corrected for the reduced permeability of gram-positive bacteria when formaldehyde was used as a fixative. The set of applied bacterial class- and subclass-specific probes yielded, on average, 58.5% (± a standard deviation of 23.0%) of the corrected eubacterial detection rates, thus indicating the necessity of additional probes for studies of biofilter communities. The Xanthomonas-specific probe presented here may serve as an efficient tool for identifying potential phytopathogens. In situ hybridization proved to be a practical tool for microbiological studies of biofiltration systems. PMID:10427047

Method for detection of a few pathogenic bacteria and determination of live versus dead cells

NASA Astrophysics Data System (ADS)

Horikawa, Shin; Chen, I.-Hsuan; Du, Songtao; Liu, Yuzhe; Wikle, Howard C.; Suh, Sang-Jin; Barbaree, James M.; Chin, Bryan A.

2016-05-01

This paper presents a method for detection of a few pathogenic bacteria and determination of live versus dead cells. The method combines wireless phage-coated magnetoelastic (ME) biosensors and a surface-scanning dectector, enabling real-time monitoring of the growth of specific bacteria in a nutrient broth. The ME biosensor used in this investigation is composed of a strip-shaped ME resonator upon which an engineered bacteriophage is coated to capture a pathogen of interest. E2 phage with high binding affinity for Salmonella Typhimurium was used as a model study. The specificity of E2 phage has been reported to be 1 in 105 background bacteria. The phage-coated ME biosensors were first exposed to a low-concentration Salmonella suspension to capture roughly 300 cells on the sensor surface. When the growth of Salmonella in the broth occurs, the mass of the biosensor increases, which results in a decrease in the biosensor's resonant frequency. Monitoring of this mass- induced resonant frequency change allows for real-time detection of the presence of Salmonella. Detection of a few bacteria is also possible by growing them to a sufficient number. The surface-scanning detector was used to measure resonant frequency changes of 25 biosensors sequentially in an automated manner as a function of time. This methodology offers direct, real-time detection, quantification, and viability determination of specific bacteria. The rate of the sensor's resonant frequency change was found to be largely dependent on the number of initially bound cells and the efficiency of cell growth.

Trends and comparison of water quality and bottom material of northeastern Arkansas streams, 1974-85, and effects of planned diversions

DOE Office of Scientific and Technical Information (OSTI.GOV)

Petersen, J.C.

1990-01-01

Water quality of several rivers in Arkansas was compared using median values at individual water quality stations. Differences were detected in several properties, including common dissolved constituents, alkalinity, nutrients, fecal coliform bacteria, trace metals, pesticides, and sediment. In bottom material, organochlorine pesticides were detected much more frequently than organophosphorus pesticides and were higher in rivers with beds of fine-gradient particles. Time trends were examined using the Seasonal Kendall test. Trends in conductance, sodium adsorption ratio, chloride, phosphorus, and ammonia were usually not detectable. Sulfate concentrations were increasing at approximately one-half of the stations studied while fecal-coliform bacteria concentrations decreased atmore » approximately one-half of the stations. The most potentially detrimental effects upon water quality resulting from surface water diversions were related to increases of common dissolved constituents. From available data, the largest of these increases would be caused by diversion from the Arkansas River. Potential effects not specifically examined include resuspension of bottom materials resulting from construction and operation of the diversion system. Use of some surface waters for artificial recharge of the alluvial aquifer may adversely affect the recharge systems or the aquifer. Possible effects include plugging of the injection well and the aquifer.« less

Detection of Bacteria Using Inkjet-Printed Enzymatic Test Strips

PubMed Central

2015-01-01

Low-cost diagnostics for drinking water contamination have the potential to save millions of lives. We report a method that uses inkjet printing to copattern an enzyme–nanoparticle sensor and substrate on a paper-based test strip for rapid detection of bacteria. A colorimetric response is generated on the paper substrate that allows visual detection of contamination without the need for expensive instrumentation. These strips demonstrate a viable nanomanufacturing strategy for low-cost bacterial detection. PMID:25318086

Classification of Salmonella Enterica serotypes with selective bands using visible/NIR hyperspectral imaging

USDA-ARS?s Scientific Manuscript database

Optical detection of foodborne bacteria such as Salmonella classifies bacteria by analyzing spectral data, and has potential for rapid detection. In this experiment hyperspectral microscopy is explored as a means for classifying five Salmonella serotypes. Initially, the microscope collects 89 spect...

Bioprobes Based on Aptamer and Silica Fluorescent Nanoparticles for Bacteria Salmonella typhimurium Detection

NASA Astrophysics Data System (ADS)

Wang, Qiu-Yue; Kang, Yan-Jun

2016-03-01

In this study, we have developed an efficient method based on single-stranded DNA (ssDNA) aptamers along with silica fluorescence nanoparticles for bacteria Salmonella typhimurium detection. Carboxyl-modified Tris(2,2'-bipyridyl)dichlororuthenium(II) hexahydrate (RuBPY)-doped silica nanoparticles (COOH-FSiNPs) were prepared using reverse microemulsion method, and the streptavidin was conjugated to the surface of the prepared COOH-FSiNPs. The bacteria S. typhimurium was incubated with a specific ssDNA biotin-labeled aptamer, and then the aptamer-bacteria conjugates were treated with the synthetic streptavidin-conjugated silica fluorescence nanoprobes (SA-FSiNPs). The results under fluorescence microscopy show that SA-FSiNPs can be applied effectively for the labeling of bacteria S. typhimurium with great photostable property. To further verify the specificity of SA-FSiNPs out of multiple bacterial conditions, variant concentrations of bacteria mixtures composed of bacteria S. typhimurium, Escherichia coli, and Bacillus subtilis were treated with SA-FSiNPs.

Bacterial community structures in air conditioners installed in Japanese residential buildings.

PubMed

Hatayama, Kouta; Oikawa, Yurika; Ito, Hiroyuki

2018-01-01

The bacterial community structures in four Japanese split-type air conditioners were analyzed using a next-generation sequencer. A variety of bacteria were detected in the air filter of an air conditioner installed on the first floor. In the evaporator of this air conditioner, bacteria belonging to the genus Methylobacterium, or the family of Sphingomonadaceae, were predominantly detected. On the other hand, the majority of bacteria detected in the air filters and evaporators of air conditioners installed on the fifth and twelfth floors belonged to the family Enterobacteriaceae. The source of bacteria belonging to the family Enterobacteriaceae may have been aerosols generated by toilet flushing in the buildings. Our results suggested the possibility that the bacterial contamination in the air conditioners was affected by the floor level on which they were installed. The air conditioner installed on the lower floor, near the ground, may have been contaminated by a variety of outdoor bacteria, whereas the air conditioners installed on floors more distant from the ground may have been less contaminated by outdoor bacteria. However, these suppositions may apply only to the specific split-type air conditioners that we analyzed, because our sample size was small.

Prevalence of pathogenic bacteria in Ixodes ricinus ticks in Central Bohemia.

PubMed

Klubal, Radek; Kopecky, Jan; Nesvorna, Marta; Sparagano, Olivier A E; Thomayerova, Jana; Hubert, Jan

2016-01-01

Bacteria associated with the tick Ixodes ricinus were assessed in specimens unattached or attached to the skin of cats, dogs and humans, collected in the Czech Republic. The bacteria were detected by PCR in 97 of 142 pooled samples including 204 ticks, i.e. 1-7 ticks per sample, collected at the same time from one host. A fragment of the bacterial 16S rRNA gene was amplified, cloned and sequenced from 32 randomly selected samples. The most frequent sequences were those related to Candidatus Midichloria midichlori (71% of cloned sequences), followed by Diplorickettsia (13%), Spiroplasma (3%), Rickettsia (3%), Pasteurella (3%), Morganella (3%), Pseudomonas (2%), Bacillus (1%), Methylobacterium (1%) and Phyllobacterium (1%). The phylogenetic analysis of Spiroplasma 16S rRNA gene sequences showed two groups related to Spiroplasma eriocheiris and Spiroplasma melliferum, respectively. Using group-specific primers, the following potentially pathogenic bacteria were detected: Borellia (in 20% of the 142 samples), Rickettsia (12%), Spiroplasma (5%), Diplorickettsia (5%) and Anaplasma (2%). In total, 68% of I. ricinus samples (97/142) contained detectable bacteria and 13% contained two or more putative pathogenic groups. The prevalence of tick-borne bacteria was similar to the observations in other European countries.

A New Primer to Amplify pmoA Gene From NC10 Bacteria in the Sediments of Dongchang Lake and Dongping Lake.

PubMed

Wang, Shenghui; Liu, Yanjun; Liu, Guofu; Huang, Yaru; Zhou, Yu

2017-08-01

Nitrite-dependent anaerobic methane oxidation (n-damo) is catalyzed by the NC10 phylum bacterium "Candidatus Methylomirabilis oxyfera" (M. oxyfera). Generally, the pmoA gene is applied as a functional marker to test and identify NC10-like bacteria. However, it is difficult to detect the NC10 bacteria from sediments of freshwater lake (Dongchang Lake and Dongping Lake) with the previous pmoA gene primer sets. In this work, a new primer cmo208 was designed and used to amplify pmoA gene of NC10-like bacteria. A newly nested PCR approach was performed using the new primer cmo208 and the previous primers cmo182, cmo682, and cmo568 to detect the NC10 bacteria. The obtained pmoA gene sequences exhibited 85-92% nucleotide identity and 95-97% amino acid sequence identity to pmoA gene of M. oxyfera. The obtained diversity of pmoA gene sequences coincided well with the diversity of 16S rRNA sequences. These results indicated that the newly designed pmoA primer cmo208 could give one more option to detect NC10 bacteria from different environmental samples.

Impact of manure fertilization on the abundance of antibiotic-resistant bacteria and frequency of detection of antibiotic resistance genes in soil and on vegetables at harvest.

PubMed

Marti, Romain; Scott, Andrew; Tien, Yuan-Ching; Murray, Roger; Sabourin, Lyne; Zhang, Yun; Topp, Edward

2013-09-01

Consumption of vegetables represents a route of direct human exposure to bacteria found in soil. The present study evaluated the complement of bacteria resistant to various antibiotics on vegetables often eaten raw (tomato, cucumber, pepper, carrot, radish, lettuce) and how this might vary with growth in soil fertilized inorganically or with dairy or swine manure. Vegetables were sown into field plots immediately following fertilization and harvested when of marketable quality. Vegetable and soil samples were evaluated for viable antibiotic-resistant bacteria by plate count on Chromocult medium supplemented with antibiotics at clinical breakpoint concentrations. DNA was extracted from soil and vegetables and evaluated by PCR for the presence of 46 gene targets associated with plasmid incompatibility groups, integrons, or antibiotic resistance genes. Soil receiving manure was enriched in antibiotic-resistant bacteria and various antibiotic resistance determinants. There was no coherent corresponding increase in the abundance of antibiotic-resistant bacteria enumerated from any vegetable grown in manure-fertilized soil. Numerous antibiotic resistance determinants were detected in DNA extracted from vegetables grown in unmanured soil. A smaller number of determinants were additionally detected on vegetables grown only in manured and not in unmanured soil. Overall, consumption of raw vegetables represents a route of human exposure to antibiotic-resistant bacteria and resistance determinants naturally present in soil. However, the detection of some determinants on vegetables grown only in freshly manured soil reinforces the advisability of pretreating manure through composting or other stabilization processes or mandating offset times between manuring and harvesting vegetables for human consumption.

Limited retention of micro-organisms using commercialized needle filters.

PubMed

Elbaz, W; McCarthy, G; Mawhinney, T; Goldsmith, C E; Moore, J E

2015-03-01

A study was undertaken to compare a commercialized needle filter with a 0.2-μm filtered epidural set and a non-filtered standard needle. No culturable bacteria were detected following filtration through the 0.2-μm filter. Bacterial breakthrough was observed with the filtered needle (pore size 5 μm) and the non-filtered needle. Filtered systems (0.2 μm) should be employed to achieve total bacterial retention. This highlights that filtration systems with different pore sizes will have varying ability to retain bacteria. Healthcare professionals need to know what type/capability of filter is implied on labels used by manufacturers, and to assess whether the specification has the desired functionality to prevent bacterial translocation through needles. Copyright © 2015 The Healthcare Infection Society. Published by Elsevier Ltd. All rights reserved.

The Hawaiian bobtail squid as a model system for selective particle capture in microfluidic systems.

NASA Astrophysics Data System (ADS)

Nawroth, Janna; McFall-Ngai, Margaret; Dabiri, John

2013-11-01

Juvenile Hawaiian bobtail squids reliably capture and isolate a single species of bacteria, Vibrio fischeri, from inhaled coastal water containing a huge background of living and non-living particles of comparable size. Biochemical mechanisms orchestrate a chain of specific interactions as soon as V.fischeri attach to the squid's internal light organ. It remains unclear, however, how the bacteria carried by the squid's ventilation currents are initially attracted to the light organ's surface. Here we present preliminary experimental data showing how arrangement and coordination of the cilia covering the light organ create a 3D flow field that facilitates advection, sieving and selective retention of flow-borne particles. These studies may inspire novel microfluidic tools for detection and capture of specific cells and particles.

R&D 100 Winner 2010: Acoustic Wave Biosensors

ScienceCinema

Larson, Richard; Branch, Darren; Edwards, Thayne

2018-01-16

The acoustic wave biosensor is innovative device that is a handheld, battery-powered, portable detection system capable of multiplex identification of a wide range of medically relevant pathogens and their biomolecular signatures — viruses, bacteria, proteins, and DNA — at clinically relevant levels. This detection occurs within minutes — not hours — at the point of care, whether that care is in a physician's office, a hospital bed, or at the scene of a biodefense or biomedical emergency.

Rapid detection of multidrug resistant Gram-negative bacilli by Cica-Beta-Test strips.

PubMed

Lavigne, J-P; Pfeiffer, C; Vidal, L; Sotto, A

2011-02-01

This study aimed to evaluate the sensitivity and specificity of a new system (Cica-Beta-Test, Kanto Chemical, Japan) for rapid detection of AmpC-derepressed, extended-spectrum β-lactamases (ESβL) and metallo-β-lactamases (MβL). Two hundred Multi-Drug Resistant (MDR)-Gram-negative bacilli were studied: 170 Enterobacteriaceae and 30 Gram-negative non-fermentative bacteria. One hundred and eighteen strains produced an ESβL, seven MβL and 75 derepressed cephalosporinases. One drop of substrate was dispensed onto the filter pad of the Cica-Beta-Test strip. The bacterial colonies were spread on the filter pad of strip. The reading was performed after 2 to 15 min: turning chromogenic indicated the positive test. Three tests were used: Cica-Beta I for detection of MDR bacteria; Cica-Beta MβL for detection of MβL-producing bacteria and Cica-Beta CVA, which distinguish ESβL and AmpC-derepressed producers. Results were compared with molecular assays. Cica-Beta-Test I has detected 194 MDR (sensitivity 97%), Cica-Beta-Test MβL has shown the presence of six MβL tested (sensitivity 85.7%). Five strains were non-MβL false positive (specificity 97.3%). Cica-Beta-Test CVA allowed the differentiation of ESβL-producing strains (109/115) and AmpC-derepressed strains (56/67) (sensitivity 94.8%, specificity 83.8%). Because of their epidemic nature, the MDR strains are screened and require strict hygienic measures patients. The simultaneous use of three strips can quickly determine the presence of MDR including ESβL and MβL. Rapid screening of MDR avoids transmission and limits the use of broad-spectrum antibiotics. Copyright © 2010 Elsevier Masson SAS. All rights reserved.

Colonization sites in carriers of ESBL-producing Gram-negative bacteria.

PubMed

van Prehn, Joffrey; Kaiser, Anna M; van der Werff, Suzanne D; van Mansfeld, Rosa; Vandenbroucke-Grauls, Christina M J E

2018-01-01

The distribution of Extended-Spectrum Beta-Lactamase-producing Gram-negative bacteria (ESBL-GNB) colonization sites is relevant for infection control guidelines on detection and follow-up of colonization. We questioned whether it is possible to rely solely on rectal swab culture for follow-up of ESBL-GNB colonization. We retrospectively assessed ESBL-GNB colonization sites in patients in a tertiary hospital in the Netherlands. The Laboratory Information Management System was queried for all bacterial cultures obtained between January 2012 and August 2016. All patients with one or more cultures positive for ESBL-GNB were identified and the distribution of ESBL-GNB positive sample sites was assessed. A subgroup analysis was performed on patients for whom at least one rectal swab specimen was available. We identified 1011 ESBL-GNB carriers with 16,578 specimens for analysis. ESBL-GNB were most frequently isolated from the rectum (506/1011), followed by the urogenital (414/1011) and respiratory tract (142/1011), and pus (136/1011). For 588 patients at least one rectal swab specimen was available. In this subgroup, ESBL-GNB colonization was detected only in the rectum in 55.4% (326/588) of patients, in 30.6% (180/588) in the rectum and a different culture site, and in 13.9% (82/588) no rectal colonization was detected. Rectal colonization with ESBL-GNB was detected in 86% of ESBL-GNB carriers. However, in 14% of ESBL-GNB carriers we did not detect rectal colonization. Therefore, samples taken for follow-up of colonization with multi-drug resistant Gram-negative bacteria (MDR-GNB) should ideally also include samples from the site where the MDR-GNB was initially found.

High sensitive virus and bacteria detection using plasma-surface-functionalized and antibody-integrated carbon nanomaterials

NASA Astrophysics Data System (ADS)

Nagatsu, Masaaki

2015-09-01

In this study we will present our recent results on the virus and bacteria detection system using the surface-functionalized carbon-encapsulated magnetic nanoparticles (NPs) fabricated by dc arc discharge, and carbon nanotube(CNT) dot-array prepared with a combined thermal and plasma CVD system. Surface functionalization of their surfaces has been carried out by plasma chemical modification using a low-pressure RF plasma for carbon-encapsulated magnetic NPs, and an ultrafine atmospheric pressure plasma jet(APPJ) for CNT dot-array substrate. After immobilization of the relevant biomolecules onto the surface of nano-structured materials, we have carried out the experiments on virus or bacteria detection using these surface-functionalized nano-structured materials. From the preliminary experiments with carbon-encapsulated magnetic NPs, we confirmed that influenza A (H1N1) virus concentration of 17.3-fold was achieved by using anti-influenza A virus hemagglutinin (HA) antibody. We have also confirmed a rapid and sensitive detection of Salmonella using the proposed method. The feasibility of CNT dot-array as a microarray biosensor has been studied by maskless functionalization of amino (-NH2) and carboxyl (-COOH) groups onto CNTs by using a ultrafine APPJ with a micro-capillary. The experimental results of chemical derivatization with the fluorescent dye showed that the CNT dot-array was not only functionalized with amino group and carboxyl group, but was also functionalized without any interference between functional groups. The success of maskless functionalization in the line pattern provides a feasibility of a multi-functionalization CNT dot-array device for future application of a microarray biosensor. This work has been supported in part by Grant-in-Aid for Scientific Research (Nos. 21110010 and 25246029) from the JSPS and the International Research Collaboration and Scientific Publication Grant (DIPA-23.04.1.673453/2015) from DGHE Indonesia.

NASA Tech Briefs, April 2009

NASA Technical Reports Server (NTRS)

2009-01-01

Topics covered include: Direct-Solve Image-Based Wavefront Sensing; Use of UV Sources for Detection and Identification of Explosives; Using Fluorescent Viruses for Detecting Bacteria in Water; Gradiometer Using Middle Loops as Sensing Elements in a Low-Field SQUID MRI System; Volcano Monitor: Autonomous Triggering of In-Situ Sensors; Wireless Fluid-Level Sensors for Harsh Environments; Interference-Detection Module in a Digital Radar Receiver; Modal Vibration Analysis of Large Castings; Structural/Radiation-Shielding Epoxies; Integrated Multilayer Insulation; Apparatus for Screening Multiple Oxygen-Reduction Catalysts; Determining Aliasing in Isolated Signal Conditioning Modules; Composite Bipolar Plate for Unitized Fuel Cell/Electrolyzer Systems; Spectrum Analyzers Incorporating Tunable WGM Resonators; Quantum-Well Thermophotovoltaic Cells; Bounded-Angle Iterative Decoding of LDPC Codes; Conversion from Tree to Graph Representation of Requirements; Parallel Hybrid Vehicle Optimal Storage System; and Anaerobic Digestion in a Flooded Densified Leachbed.

Utilization of an Airborne Plant Chlorophyll Imaging System for Detection of Septic System Malfunction

NASA Technical Reports Server (NTRS)

Spiering, Bruce A.; Carter, Gregory A.

2001-01-01

Malfunctioning, or leaking, sewer systems increase the supply of water and nutrients to surface vegetation. Excess nutrients and harmful bacteria in the effluent pollute ground water and local water bodies and are dangerous to humans and the aquatic ecosystems. An airborne multispectral plant chlorophyll imaging system (PCIS) was used to identify growth patterns in the vegetation covering onsite and public sewer systems. The objective was to evaluate overall performance of the PCIS as well as to determine the best operational configuration for this application. The imaging system was flown in a light aircraft over selected locations Mobile County, Alabama. Calibration panels were used to help characterize instrument performance. Results demonstrated that the PCIS performed well and was capable of detecting septic leakage patterns from altitudes as high as 915 m. From 915 m, 6 of 18 sites were suspected to have sewage leakage. Subsequent ground inspections confirmed leakage on 3 of the 6 sites. From 610 m, 3 of 8 known leakage sites were detected. Tree cover and shadows near residential structures prevented detection of several known malfunctioning systems. Also some leakages known to occur in clear areas were not detected. False detections occurred in areas characterized by surface water drainage problems or recent excavation.

Probiotic culture survival and implications in fermented frozen yogurt characteristics.

PubMed

Davidson, R H; Duncan, S E; Hackney, C R; Eigel, W N; Boling, J W

2000-04-01

Low-fat ice cream mix was fermented with probiotic-supplemented and traditional starter culture systems and evaluated for culture survival, composition, and sensory characteristics of frozen product. Fermentations were stopped when the titratable acidity reached 0.15% greater than the initial titratable acidity (end point 1) or when the pH reached 5.6 (end point 2). Mix was frozen and stored for 11 wk at -20 degrees C. The traditional yogurt culture system contained the strains Streptococcus salivarius ssp. thermophilus and Lactobacillus delbrueckii ssp. bulgaricus. The probiotic-supplemented system contained the traditional cultures as well as Bifidobacterium longum and Lactobacillus acidophilus. We compared recovery of Bifodobacterium by three methods, a repair-detection system with roll-tubes and plates on modified bifid glucose medium and plates with maltose + galactose reinforced clostridial medium. Culture bacteria in both systems did not decrease in the yogurt during frozen storage. The roll-tube method with modified bifid glucose agar and repair detection system provided at least one-half log10 cfu/ml higher recovery of B. longum compared with recoveries using modified bifid glucose agar or maltose + galactose reinforced clostridial agar on petri plates. No change in concentrations of lactose or protein for products fermented with either culture system occurred during storage. Acid flavor was more intense when product was fermented to pH 5.6, but yogurt flavor was not intensified. The presence of probiotic bacteria in the supplemented system seemed to cause no differences in protein and lactose concentration and sensory characteristics.

Characterization of airborne bacteria at an underground subway station.

PubMed

Dybwad, Marius; Granum, Per Einar; Bruheim, Per; Blatny, Janet Martha

2012-03-01

The reliable detection of airborne biological threat agents depends on several factors, including the performance criteria of the detector and its operational environment. One step in improving the detector's performance is to increase our knowledge of the biological aerosol background in potential operational environments. Subway stations are enclosed public environments, which may be regarded as potential targets for incidents involving biological threat agents. In this study, the airborne bacterial community at a subway station in Norway was characterized (concentration level, diversity, and virulence- and survival-associated properties). In addition, a SASS 3100 high-volume air sampler and a matrix-assisted laser desorption ionization-time of flight mass spectrometry-based isolate screening procedure was used for these studies. The daytime level of airborne bacteria at the station was higher than the nighttime and outdoor levels, and the relative bacterial spore number was higher in outdoor air than at the station. The bacterial content, particle concentration, and size distribution were stable within each environment throughout the study (May to September 2010). The majority of the airborne bacteria belonged to the genera Bacillus, Micrococcus, and Staphylococcus, but a total of 37 different genera were identified in the air. These results suggest that anthropogenic sources are major contributors to airborne bacteria at subway stations and that such airborne communities could harbor virulence- and survival-associated properties of potential relevance for biological detection and surveillance, as well as for public health. Our findings also contribute to the development of realistic testing and evaluation schemes for biological detection/surveillance systems by providing information that can be used to mimic real-life operational airborne environments in controlled aerosol test chambers.

Characterization of Airborne Bacteria at an Underground Subway Station

PubMed Central

Dybwad, Marius; Granum, Per Einar; Bruheim, Per

2012-01-01

The reliable detection of airborne biological threat agents depends on several factors, including the performance criteria of the detector and its operational environment. One step in improving the detector's performance is to increase our knowledge of the biological aerosol background in potential operational environments. Subway stations are enclosed public environments, which may be regarded as potential targets for incidents involving biological threat agents. In this study, the airborne bacterial community at a subway station in Norway was characterized (concentration level, diversity, and virulence- and survival-associated properties). In addition, a SASS 3100 high-volume air sampler and a matrix-assisted laser desorption ionization–time of flight mass spectrometry-based isolate screening procedure was used for these studies. The daytime level of airborne bacteria at the station was higher than the nighttime and outdoor levels, and the relative bacterial spore number was higher in outdoor air than at the station. The bacterial content, particle concentration, and size distribution were stable within each environment throughout the study (May to September 2010). The majority of the airborne bacteria belonged to the genera Bacillus, Micrococcus, and Staphylococcus, but a total of 37 different genera were identified in the air. These results suggest that anthropogenic sources are major contributors to airborne bacteria at subway stations and that such airborne communities could harbor virulence- and survival-associated properties of potential relevance for biological detection and surveillance, as well as for public health. Our findings also contribute to the development of realistic testing and evaluation schemes for biological detection/surveillance systems by providing information that can be used to mimic real-life operational airborne environments in controlled aerosol test chambers. PMID:22247150

Small Talk: Cell-to-Cell Communication in Bacteria

ScienceCinema

Bassler, Bonnie [Princeton University, Princeton, New Jersey, United States

2017-12-09

Cell-cell communication in bacteria involves the production, release, and subsequent detection of chemical signaling molecules called autoinducers. This process, called quorum sensing, allows bacteria to regulate gene expression on a population-wide scale. Processes controlled by quorum sensing are usually ones that are unproductive when undertaken by an individual bacterium but become effective when undertaken by the group. For example, quorum sensing controls bioluminescence, secretion of virulence factors, biofilm formation, sporulation, and the exchange of DNA. Thus, quorum sensing is a mechanism that allows bacteria to function as multi-cellular organisms. Bacteria make, detect, and integrate information from multiple autoinducers, some of which are used exclusively for intra-species communication while others enable communication between species. Research is now focused on the development of therapies that interfere with quorum sensing to control bacterial virulence.

Identification of feces by detection of Bacteroides genes.

PubMed

Nakanishi, Hiroaki; Shojo, Hideki; Ohmori, Takeshi; Hara, Masaaki; Takada, Aya; Adachi, Noboru; Saito, Kazuyuki

2013-01-01

In forensic science, the identification of feces is very important in a variety of crime investigations. However, no sensitive and simple fecal identification method using molecular biological techniques has been reported. Here, we focused on the fecal bacteria, Bacteroides uniformis, Bacteroides vulgatus and Bacteroides thetaiotaomicron, and developed a novel fecal identification method by detection of the gene sequences specific to these bacteria in various body (feces, blood, saliva, semen, urine, vaginal fluids and skin surfaces) and forensic (anal adhesions) specimens. Bacterial gene detection was performed by real-time PCR using a minor groove binding probe to amplify the RNA polymerase β-subunit gene of B. uniformis and B. vulgatus, and the α-1-6 mannanase gene of B. thetaiotaomicron. At least one of these bacteria was detected in the feces of 20 donors; the proportions of B. uniformis, B. vulgatus and B. thetaiotaomicron were 95, 85 and 60%, respectively. Bacteroides vulgatus was also detected in one of six vaginal fluid samples, but B. thetaiotaomicron and B. uniformis were not detected in body samples other than feces. Further, we applied this method to forensic specimens from 18 donors. Eighteen anal adhesions also contained at least one of three bacteria; B. uniformis, B. vulgatus and B. thetaiotaomicron were detected in 89, 78 and 56%, respectively, of the specimens. Thus, these bacteria were present at a high frequency in the fecal and forensic specimens, while either B. uniformis or B. vulgatus was detected in all samples. Therefore, B. uniformis and B. vulgatus represent more appropriate target species than B. thetaiotaomicron for the identification of fecal material. If B. vulgatus and/or B. uniformis are detected, it is likely that the sample contains feces. Taken together, our results suggest that the use of molecular biological techniques will aid the detection of feces in forensic practice, although it is possible that the samples contained both feces and vaginal fluid. Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.

Staphylococcus aureus detection in blood samples by silica nanoparticle-oligonucleotides conjugates.

PubMed

Borsa, Baris A; Tuna, Bilge G; Hernandez, Frank J; Hernandez, Luiza I; Bayramoglu, Gulay; Arica, M Yakup; Ozalp, V Cengiz

2016-12-15

A fast, specific and sensitive homogeneous assay for Staphylococcus aureus detection was developed by measuring the activity of secreted nuclease from the bacteria via a modified DNA oligonucleotide. As biosensor format, an effective system, Nanokeepers as previously reported, were used for triggered release of confined fluorophores, and hence specific detection of S. aureus on nuclease activity was obtained. The interference from blood components for fluorescent quantification was eliminated by a pre-purification by aptamer-functionalized silica magnetic nanoparticles. The reported assay system was exclusively formed by nucleic acid oligos and magnetic or mesoporous silica nanoparticles, that can be used on blood samples in a stepwise manner. The assay was successfully used as a sensing platform for the specific detection of S. aureus cells as low as 682 CFU in whole blood. Copyright © 2016 Elsevier B.V. All rights reserved.

Ancient acquisition of "alginate utilization loci" by human gut microbiota.

PubMed

Mathieu, Sophie; Touvrey-Loiodice, Mélanie; Poulet, Laurent; Drouillard, Sophie; Vincentelli, Renaud; Henrissat, Bernard; Skjåk-Bræk, Gudmund; Helbert, William

2018-05-23

In bacteria from the phylum Bacteroidetes, the genes coding for enzymes involved in polysaccharide degradation are often colocalized and coregulated in so-called "polysaccharide utilization loci" (PULs). PULs dedicated to the degradation of marine polysaccharides (e.g. laminaran, ulvan, alginate and porphyran) have been characterized in marine bacteria. Interestingly, the gut microbiome of Japanese individuals acquired, by lateral transfer from marine bacteria, the genes involved in the breakdown of porphyran, the cell wall polysaccharide of the red seaweed used in maki. Sequence similarity analyses predict that the human gut microbiome also encodes enzymes for the degradation of alginate, the main cell wall polysaccharide of brown algae. We undertook the functional characterization of diverse polysaccharide lyases from family PL17, frequently found in marine bacteria as well as those of human gut bacteria. We demonstrate here that this family is polyspecific. Our phylogenetic analysis of family PL17 reveals that all alginate lyases, which have all the same specificity and mode of action, cluster together in a very distinct subfamily. The alginate lyases found in human gut bacteria group together in a single clade which is rooted deeply in the PL17 tree. These enzymes were found in PULs containing PL6 enzymes, which also clustered together in the phylogenetic tree of PL6. Together, biochemical and bioinformatics analyses suggest that acquisition of this system appears ancient and, because only traces of two successful transfers were detected upon inspection of PL6 and PL17 families, the pace of acquisition of marine polysaccharide degradation system is probably very slow.

A light-up probe with aggregation-induced emission characteristics (AIE) for selective imaging, naked-eye detection and photodynamic killing of Gram-positive bacteria.

PubMed

Feng, Guangxue; Yuan, Youyong; Fang, Hu; Zhang, Ruoyu; Xing, Bengang; Zhang, Guanxin; Zhang, Deqing; Liu, Bin

2015-08-11

We report the design and synthesis of a red fluorescent AIE light-up probe for selective recognition, naked-eye detection, and image-guided photodynamic killing of Gram-positive bacteria, including vancomycin-resistant Enterococcus strains.

Capillary isoelectric focusing of probiotic bacteria from cow's milk in tapered fused silica capillary with off-line matrix-assisted laser desorption/ionization time-of-flight mass spectrometry identification.

PubMed

Horká, Marie; Karásek, Pavel; Salplachta, Jiří; Růžička, Filip; Vykydalová, Marie; Kubesová, Anna; Dráb, Vladimír; Roth, Michal; Slais, Karel

2013-07-25

In this study, combination of capillary isoelectric focusing (CIEF) in tapered fused silica (FS) capillary with matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) is presented as an efficient approach for unambiguous identification of probiotic bacteria in real sample. For this purpose, bacteria within genus Lactobacillus were selected as model bioanalytes and cow's milk was selected as a biological sample. CIEF analysis of both the cultivated bacteria and the bacteria in the milk was optimized and isoelectric points characterizing the examined bacteria were subsequently determined independently of the bacterial sample origin. The use of tapered FS capillary significantly enhanced the separation capacity and efficiency of the CIEF analyses performed. In addition, the cell number injected into the tapered FS capillary was quantified and an excellent linearity of the calibration curves was achieved which enabled quantitative analysis of the bacteria by CIEF with UV detection. The minimum detectable number of bacterial cells was 2×10(6) mL(-1). Finally, cow's milk spiked with the selected bacterium was analyzed by CIEF in tapered FS capillary, the focused and detected bacterial cells were collected from the capillary, deposited onto the cultivation medium, and identified using MALDI-TOF MS afterward. Our results have revealed that the proposed procedure can be advantageously used for unambiguous identification of probiotic bacteria in a real sample. Copyright © 2013 Elsevier B.V. All rights reserved.

Centrifugal sedimentation immunoassays for multiplexed detection of enteric bacteria in ground water

DOE PAGES

Litvinov, Julia; Moen, Scott T.; Koh, Chung-Yan; ...

2016-01-01

Water-born pathogens pose significant threat to the global population and early detection plays an important role both in making drinking water safe, as well as in diagnostics and treatment of water-borne diseases. We present an innovative centrifugal microfluidic platform (SpinDx) for detection of bacterial pathogens using bead-based immunoassays. Our approach is based on binding of pathogens to antibody-functionalized capture particles followed by sedimentation of the particles through a density-media in a microfluidic disk and quantification by fluorescence microscopy. Our platform is fast (20 min), sensitive (10 3 CFU/mL), requires minimal sample preparation, and can detect multiple pathogens simultaneously with sensitivitymore » similar to that required by the EPA. We demonstrate detection of a panel of enteric bacteria (Escherichia coli, Salmonella typhimurium, Shigella, Listeria, and Campylobacter) at concentrations as low as 10 3 CFU/mL or 30 bacteria per reaction.« less

Electrical response of culture media during bacterial growth on a paper-based device

NASA Astrophysics Data System (ADS)

Srimongkon, Tithimanan; Buerkle, Marius; Nakamura, Akira; Enomae, Toshiharu; Ushijima, Hirobumi; Fukuda, Nobuko

2017-05-01

In this work, we evaluated the feasibility of a paper-based bacterial detection system. The paper served as a substrate for the measurement electrodes and the culture medium. Using a printing technique, we patterned gold electrodes onto the paper substrate and applied Luria broth (LB) agar gel as a culture medium on top of the electrodes. As the first step towards the development of a bacterial detection system, we determined changes in the surface potential during bacterial growth and monitored these changes over 24 h. This allowed us to correlate changes in the surface potential with the different growth phases of the bacteria.

Quantification of carious pathogens in the interdental microbiota of young caries-free adults.

PubMed

Bourgeois, Denis; David, Alexandra; Inquimbert, Camille; Tramini, Paul; Molinari, Nicolas; Carrouel, Florence

2017-01-01

The majority of caries lesions in adults occur on the proximal tooth surfaces of the posterior teeth. A comprehensive study of the composition of the oral microbiota is fundamental for a better understanding of the etiology of interdental caries. Twenty-five caries-free subjects (20-35 years old) were enrolled in the study. The interdental biofilm of four interdental sites were collected. The real-time polymerase chain reaction (PCR) methodology were used to quantify (i) the following bacteria: Streptococcus spp., Streptococcus mutans, Lactobacillus spp., Enterococcus spp., and Enterococcus faecalis; (ii) the fungus Candida albicans; and (iii) total bacteria. Streptococcus spp. was the most abundant species, followed by Lactobacillus spp. and Enterococcus spp. Streptococcus spp. and Lactobacillus spp. were detected at all tested sites and Enterococcus spp. at 99% of sites. S. mutans was detected at only 28% of the tested sites and C. albicans was detected at 11% of sites. E. faecalis was never detected. In 54.5% of the biofilm inhabited by C. albicans, S. mutans was present. Moreover, 28% of the ID sites co-expressed S. mutans and Lactobacillus spp. The studied pathogens were organized into two correlated groups of species. Strikingly, the fungus C. albicans and the bacteria Enterococcus spp. cluster together, whereas Streptococcus spp., S. mutans and Lactobacillus spp. form one distinct cluster. The interdental biofilm of young caries-free adults is comprised of pathogens that are able to induce interproximal caries. That several of these pathogens are implicated in heart disease or other systemic diseases is an argument for the disruption of interdental biofilms using daily oral hygiene.

Detection of Rickettsia and Anaplasma from hard ticks in Thailand.

PubMed

Malaisri, Premnika; Hirunkanokpun, Supanee; Baimai, Visut; Trinachartvanit, Wachareeporn; Ahantarig, Arunee

2015-12-01

We collected a total of 169 adult hard ticks and 120 nymphs from under the leaves of plants located along tourist nature trails in ten localities. The results present data examining the vector competence of ticks of different genera and the presence of Rickettsia and Anaplasma species. The ticks belonged to three genera, Amblyomma, Dermacentor, and Haemaphysalis, comprising 11 species. Rickettsia bacteria were detected at three collection sites, while Anaplasma bacteria were detected at only one site. Phylogenetic analysis revealed new rickettsia genotypes from Thailand that were closely related to Rickettsia tamurae, Rickettsia monacensis, and Rickettsia montana. This study was also the first to show that Anaplasma bacteria are found in Haemaphysalis shimoga ticks and are closely related evolutionarily to Anaplasma bovis. These results provide additional information for the geographical distribution of tick species and tick-borne bacteria in Thailand and can therefore be applied for ecotourism management. © 2015 The Society for Vector Ecology.

Detection and identification of intestinal pathogenic bacteria by hybridization to oligonucleotide microarrays

PubMed Central

Jin, Lian-Qun; Li, Jun-Wen; Wang, Sheng-Qi; Chao, Fu-Huan; Wang, Xin-Wei; Yuan, Zheng-Quan

2005-01-01

AIM: To detect the common intestinal pathogenic bacteria quickly and accurately. METHODS: A rapid (<3 h) experimental procedure was set up based upon the gene chip technology. Target genes were amplified and hybridized by oligonucleotide microarrays. RESULTS: One hundred and seventy strains of bacteria in pure culture belonging to 11 genera were successfully discriminated under comparatively same conditions, and a series of specific hybridization maps corresponding to each kind of bacteria were obtained. When this method was applied to 26 divided cultures, 25 (96.2%) were identified. CONCLUSION: Salmonella sp., Escherichia coli, Shigella sp., Listeria monocytogenes, Vibrio parahaemolyticus, Staphylococcus aureus, Proteus sp., Bacillus cereus, Vibrio cholerae, Enterococcus faecalis, Yersinia enterocolitica, and Campylobacter jejuni can be detected and identified by our microarrays. The accuracy, range, and discrimination power of this assay can be continually improved by adding further oligonucleotides to the arrays without any significant increase of complexity or cost. PMID:16437687

A PCR technique to detect enterotoxigenic and verotoxigenic Escherichia coli in boar semen samples.

PubMed

Bussalleu, E; Pinart, E; Yeste, M; Briz, M; Sancho, S; Torner, E; Bonet, S

2012-08-01

In semen, bacteria's isolation from a pure culture is complex, laborious and easily alterable by the presence of antibiotics and inhibitors. We developed a PCR technique to detect the presence of the enterotoxigenic (ETEC) and verotoxigenic Escherichia coli (VTEC) (strains with high prevalence in the swine industry) in semen by adapting the protocols developed by Zhang et al. (2007) and Yilmaz et al. (2006). We artificially inoculated extended semen samples at different infective concentrations of bacteria (from 10(2) to 10(8) bacteria ml(-1)) with two enterotoxigenic and verotoxigenic strains, and performed two multiplex and one conventional PCR. This technique proved to be a quick, useful and reliable tool to detect the presence of ETEC and VTEC up to an infective dose of 10(5) bacteria ml(-1) in semen. Copyright © 2011 Elsevier Ltd. All rights reserved.

Detection of Salmonella sp., Vibrio sp. and total plate count bacteria on blood cockle (Anadara granosa)

NASA Astrophysics Data System (ADS)

Ekawati, ER; Yusmiati, S. N. H.

2018-01-01

Blood cockle (Anadara granosa) has high level of zinc and protein, which is beneficial for therapeutic function for malnourished particularly stunting case in children. Zinc in animal foods is more absorbable than that from vegetable food. Blood cockle (Anadara granosa) is rich in nutrient and an excellent environment for the growth of microorganisms. This research aimed to identify the contamination of Salmonella sp., Vibrio sp. and total plate count bacteria on blood cockle (Anadara granosa). This was observation research with laboratory analysis. Salmonella sp. and Vibrio sp. were detected from blood cockle. Total plate count was determine of the total amount of the bacteria. Results detected from 20 samples of blood cockle showed that all samples were negative of Salmonella sp. and 1 sample positive Vibrio sp. The result of total plate count bacteria was < 5 x 105 colony/g sample.

Contribution of bacteria-like particles to PM2.5 aerosol in urban and rural environments

NASA Astrophysics Data System (ADS)

Wolf, R.; El-Haddad, I.; Slowik, J. G.; Dällenbach, K.; Bruns, E.; Vasilescu, J.; Baltensperger, U.; Prévôt, A. S. H.

2017-07-01

We report highly time-resolved estimates of airborne bacteria-like particle concentrations in ambient aerosol using an Aerodyne aerosol mass spectrometer (AMS). AMS measurements with a newly developed PM2.5 and the standard (PM1) aerodynamic lens were performed at an urban background site (Zurich) and at a rural site (Payerne) in Switzerland. Positive matrix factorization using the multilinear engine (ME-2) implementation was used to estimate the contribution of bacteria-like particles to non-refractory organic aerosol. The success of the method was evaluated by a size-resolved analysis of the organic mass and the analysis of single particle mass spectra, which were detected with a light scattering system integrated into the AMS. Use of the PM2.5 aerodynamic lens increased measured bacteria-like concentrations, supporting the analysis method. However, at all sites, the low concentrations of this component suggest that airborne bacteria constitute a minor fraction of non-refractory PM2.5 organic aerosol mass. Estimated average mass concentrations were below 0.1 μg/m3 and relative contributions were lower than 2% at both sites. During rainfall periods, concentrations of the bacteria-like component increased considerably reaching a short-time maximum of approximately 2 μg/m3 at the Payerne site in summer.

Role of the gut microbiota in host appetite control: bacterial growth to animal feeding behaviour.

PubMed

Fetissov, Sergueï O

2017-01-01

The life of all animals is dominated by alternating feelings of hunger and satiety - the main involuntary motivations for feeding-related behaviour. Gut bacteria depend fully on their host for providing the nutrients necessary for their growth. The intrinsic ability of bacteria to regulate their growth and to maintain their population within the gut suggests that gut bacteria can interfere with molecular pathways controlling energy balance in the host. The current model of appetite control is based mainly on gut-brain signalling and the animal's own needs to maintain energy homeostasis; an alternative model might also involve bacteria-host communications. Several bacterial components and metabolites have been shown to stimulate intestinal satiety pathways; at the same time, their production depends on bacterial growth cycles. This short-term bacterial growth-linked modulation of intestinal satiety can be coupled with long-term regulation of appetite, controlled by the neuropeptidergic circuitry in the hypothalamus. Indeed, several bacterial products are detected in the systemic circulation, which might act directly on hypothalamic neurons. This Review analyses the data relevant to possible involvement of the gut bacteria in the regulation of host appetite and proposes an integrative homeostatic model of appetite control that includes energy needs of both the host and its gut bacteria.

Effect of associated bacteria on the growth and toxicity of Alexandrium catenella.

PubMed

Uribe, Paulina; Espejo, Romilio T

2003-01-01

Saprophytic bacteria in cultures of the marine dinoflagellate Alexandrium catenella were removed to assess their effect on growth and paralytic shellfish poisoning toxin production of this dinoflagellate. The actual axenic status was demonstrated by the lack of observable bacteria both immediately after treatment and following extended incubation in the absence of antibiotics. Bacteria were measured by counting CFU and also by epifluorescence microscopy and PCR amplification of bacterial 16S-23S spacer ribosomal DNA to detect noncultivable bacteria. Removal of bacteria did not have any effect on the growth of the dinoflagellate except for the inhibition of A. catenella disintegration after reaching the stationary phase. Toxicity was determined in dinoflagellate cell extracts by different methods: high-performance liquid chromatography (HPLC); an electrophysiological test called the Electrotest, which measures the inhibition of saxitoxin-sensitive Na(+) channels expressed in a cell line; and a mouse bioassay, which measures the toxic effect on the whole mammal neuromuscular system. A lower toxicity of the dinoflagellates in axenic culture was observed by these three methods, though the difference was significant only by the mouse bioassay and HPLC methods. Altogether the results indicate that axenic cultures of A. catenella are able to produce toxin, though the total toxicity is probably diminished to about one-fifth of that in nonaxenic cultures.

Antibiotic-resistant bacteria in the Hudson River Estuary linked to wet weather sewage contamination.

PubMed

Young, Suzanne; Juhl, Andrew; O'Mullan, Gregory D

2013-06-01

Heterotrophic bacteria resistant to tetracycline and ampicillin were assessed in waterways of the New York City metropolitan area using culture-dependent approaches and 16S rRNA gene sequence analysis of resultant isolates. Resistant microbes were detected at all 10 sampling sites in monthly research cruises on the lower Hudson River Estuary (HRE), with highest concentrations detected at nearshore sites. Higher frequency sampling was conducted in Flushing Bay, to enumerate resistant microbes under both dry and wet weather conditions. Concentrations of ampicillin- and tetracycline-resistant bacteria, in paired samples, were positively correlated with one another and increased following precipitation. Counts of the fecal indicator, Enterococcus, were positively correlated with levels of resistant bacteria, suggesting a shared sewage-associated source. Analysis of 16S rRNA from isolates identified a phylogenetically diverse group of resistant bacteria, including genera containing opportunistic pathogens. The occurrence of Enterobacteriaceae, a family of enteric bacteria, was found to be significantly higher in resistant isolates compared to total heterotrophic bacteria and increased following precipitation. This study is the first to document the widespread distribution of antibiotic-resistant bacteria in the HRE and to demonstrate clearly a link between the abundance of antibiotic-resistant bacteria and levels of sewage-associated bacteria in an estuary.

Isolation, amplification and characterization of foodborne pathogen disease bacteria gene for rapid kit test development

NASA Astrophysics Data System (ADS)

Nurjayadi, M.; Santoso, I.; Kartika, I. R.; Kurniadewi, F.; Saamia, V.; Sofihan, W.; Nurkhasanah, D.

2017-07-01

There is a lot of public concern over food safety. Food-safety cases recently, including many food poisoning cases in both the developed and developing countries, considered to be the national security threats which involved police investigation. Quick and accurate detection methods are needed to handle the food poisoning cases with a big number of sufferers at the same time. Therefore, the research is aimed to develop a specific, sensitive, and rapid result molecular detection tool for foodborne pathogen bacteria. We, thus, propose genomic level approach with Polymerase Chain Reaction. The research has successfully produced a specific primer to perform amplification to fim-C S. typhi, E. coli, and pef Salmonella typhimurium genes. The electrophoresis result shows that amplification products are 95 base pairs, 121 base pairs, and 139 base pairs; and all three genes are in accordance with the size of the in silico to third genes bacteria. In conclusion, the research has been successfully designed a specific detection tool to three foodborne pathogen bacteria genes. Further stages test and the uses of Real-time PCR in the detection are still in the trial process for better detection method.

Recovery and diversity of heterotrophic bacteria from chlorinated drinking waters.

PubMed Central

Maki, J S; LaCroix, S J; Hopkins, B S; Staley, J T

1986-01-01

Heterotrophic bacteria were enumerated from the Seattle drinking water catchment basins and distribution system. The highest bacterial recoveries were obtained by using a very dilute medium containing 0.01% peptone as the primary carbon source. Other factors favoring high recovery were the use of incubation temperatures close to that of the habitat and an extended incubation (28 days or longer provided the highest counts). Total bacterial counts were determined by using acridine orange staining. With one exception, all acridine orange counts in chlorinated samples were lower than those in prechlorinated reservoir water, indicating that chlorination often reduces the number of acridine orange-detectable bacteria. Source waters had higher diversity index values than did samples examined following chlorination and storage in reservoirs. Shannon index values based upon colony morphology were in excess of 4.0 for prechlorinated source waters, whereas the values for final chlorinated tap waters were lower than 2.9. It is not known whether the reduction in diversity was due solely to chlorination or in part to other factors in the water treatment and distribution system. Based upon the results of this investigation, we provide a list of recommendations for changes in the procedures used for the enumeration of heterotrophic bacteria from drinking waters. Images PMID:3524453

Deoxyribonucleic Acid Probes Analyses for the Detection of Periodontal Pathogens.

PubMed

Al Yahfoufi, Zoubeida; Hadchiti, Wahib; Berberi, Antoine

2015-09-01

In clinical microbiology several techniques have been used to identify bacteria. Recently, Deoxyribonucleic acid (DNA)-based techniques have been introduced to detect human microbial pathogens in periodontal diseases. Deoxyribonucleic acid probes can detect bacteria at a very low level if we compared with the culture methods. These probes have shown rapid and cost-effective microbial diagnosis, good sensitivity and specificity for some periodontal pathogens in cases of severe periodontitis. Eighty-five patients were recruited for the study. Twenty-one subjects ranging between 22 and 48 years of age fulfilled the inclusion and exclusion criteria. Seventy-eight samples became available for DNA probe analysis from the deepest pockets in each quadrant. All 21 patients showed positive results for Prevotella intermedia; also, Prevotella gingivalis was identified in 19 subjects, Aggregatibacter actinomycetemcomitans in 6 subjects. P. intermedia was diagnosed positive in 82% of the subgingival samples taken, 79% for P. gingivalis, and 23% for A. actinomycetemcomitans. This study shows a high frequency of putative periodontal pathogens by using DNA probe technology, which is semi-quantitative in this study. Deoxyribonucleic acid probes can detect bacteria at very low level about 10(3) which is below the detection level of culture methods. The detection threshold of cultural methods. The three types of bacteria can be detected rapidly with high sensitivity by using the DNA probe by general practitioners, and thus can help in the diagnosis process and the treatment.

Quantification of two forms of green sulfur bacteria in their natural habitat using bacteriochlorophyll fluorescence spectra

NASA Astrophysics Data System (ADS)

Kharcheva, Anastasia V.; Zhiltsova, Anna A.; Lunina, Olga N.; Savvichev, Alexander S.; Patsaeva, Svetlana V.

2016-04-01

Detection of phototropic organisms in their natural habitat using optical instruments operating under water is urgently needed for many tasks of ecological monitoring. While fluorescence methods are widely applied nowadays to detect and characterize phytoplankton communities, the techniques for detection and recognition of anoxygenic phototrophs are considered challenging. Differentiation of the forms of anoxygenic green sulfur bacteria in natural water using spectral techniques remains problematic. Green sulfur bacteria could be found in two forms, green-colored (containing BChl d in pigment compound) and brown-colored (containing BChl e), have the special ecological niche in such reservoirs. Separate determination of these microorganisms by spectral methods is complicated because of similarity of spectral characteristics of their pigments. We describe the novel technique of quantification of two forms of green sulfur bacteria directly in water using bacteriochlorophyll fluorescence without pigment extraction. This technique is noninvasive and could be applied in remote mode in the water bodies with restricted water circulation to determine simultaneously concentrations of two forms of green sulfur bacteria in their natural habitat.

Escherichia coli Fiber Sensors Using Concentrated Dielectrophoretic Force with Optical Defocusing Method.

PubMed

Tai, Yi-Hsin; Lee, Chia-Wei; Chang, Dao-Ming; Lai, Yu-Sheng; Huang, Ding-Wei; Wei, Pei-Kuen

2018-05-25

A sensitive tapered optical fiber tip combined with dielectrophoretic (DEP) trapping was used for rapid and label-free detection of bacteria in water. The angular spectrum of the optical field at the fiber tip was changed with the surrounding refractive index (RI). By measuring far-field intensity change at the defocus plane, the intensity sensitivity was up to 95 200%/RIU (RI unit), and the detection limit was 5.2 × 10 -6 RIU at 0.5% intensity stability. By applying an AC voltage to a Ti/Al coated fiber tip and an indium-tin-oxide glass, the DEP force effectively trapped the Escherichia coli ( E. coli) near the fiber tip. Those bacteria can be directly measured from optical intensity change due to the increase of surrounding RI. By immobilizing the antibody on the Ti/Al fiber tip, the tests for specific K12 bacteria and nonspecific BL21 bacteria verified the specificity. The antibody-immobilized Ti/Al coated fiber tip with DEP trapping can detect bacteria at a concentration about 100 CFU/mL.

Simultaneous aptasensor for multiplex pathogenic bacteria detection based on multicolor upconversion nanoparticles labels.

PubMed

Wu, Shijia; Duan, Nuo; Shi, Zhao; Fang, Congcong; Wang, Zhouping

2014-03-18

A highly sensitive and specific multiplex method for the simultaneous detection of three pathogenic bacteria was fabricated using multicolor upconversion nanoparticles (UCNPs) as luminescence labels coupled with aptamers as the molecular recognition elements. Multicolor UCNPs were synthesized via doping with various rare-earth ions to obtain well-separated emission peaks. The aptamer sequences were selected using the systematic evolution of ligands by exponential enrichment (SELEX) strategy for Staphylococcus aureus, Vibrio parahemolyticus, and Salmonella typhimurium. When applied in this method, aptamers can be used for the specific recognition of the bacteria from complex mixtures, including those found in real food matrixes. Aptamers and multicolor UCNPs were employed to selectively capture and simultaneously quantify the three target bacteria on the basis of the independent peaks. Under optimal conditions, the correlation between the concentration of three bacteria and the luminescence signal was found to be linear from 50-10(6) cfu mL(-1). Improved by the magnetic separation and concentration effect of Fe3O4 magnetic nanoparticles, the limits of detection of the developed method were found to be 25, 10, and 15 cfu mL(-1) for S. aureus, V. parahemolyticus, and S. typhimurium, respectively. The capability of the bioassay in real food samples was also investigated, and the results were consistent with experimental results obtained from plate-counting methods. This proposed method for the detection of various pathogenic bacteria based on multicolor UCNPs has great potential in the application of food safety and multiplex nanosensors.

The Detection Method of Escherichia coli in Water Resources: A Review

NASA Astrophysics Data System (ADS)

Nurliyana, M. R.; Sahdan, M. Z.; Wibowo, K. M.; Muslihati, A.; Saim, H.; Ahmad, S. A.; Sari, Y.; Mansor, Z.

2018-04-01

This article reviews several approaches for Escherichia coli (E. coli) bacteria detection from conventional methods, emerging method and goes to biosensor-based techniques. Detection and enumeration of E. coli bacteria usually required long duration of time in obtaining the result since laboratory-based approach is normally used in its assessment. It requires 24 hours to 72 hours after sampling to process the culturing samples before results are available. Although faster technique for detecting E. coli in water such as Polymerase Chain Reaction (PCR) and Enzyme-Linked Immunosorbent Assay (ELISA) have been developed, it still required transporting the samples from water resources to the laboratory, high-cost, complicated equipment usage, complex procedures, as well as the requirement of skilled specialist to cope with the complexity which limit their wide spread practice in water quality detection. Recently, development of biosensor device that is easy to perform, portable, highly sensitive and selective becomes indispensable in detecting extremely lower consolidation of pathogenic E. coli bacteria in water samples.

The Novel Immunobiosensors for Detection of Escherichia coli O157:H7 Using Electrochemical Impedance Spectroscopy.

PubMed

Zhang, Deng; Chen, Songyue; Qin, Lifeng; Li, Rong; Wang, Ping; Li, Yanbin

2005-01-01

Immunobiosensors were developed for detection of Escherichia coli O157:H7 based on the surface immobilization of monoclone antibodies onto indium tin oxide (ITO) electrodes. The immobilization of antibodies onto ITO chips was carried out by silanization. The effects of epoxysilane monolayer, the antibody layer on the electrochemical properties of the electrode, and the combined target bacteria were analyzed through cyclic voltammetry and electrochemical impedance spectroscopy. By using Randles model as the equivalent circuit, the concentration of the target bacteria can be quantitatively analyzed in terms of the change of electron transfer resistance. The biosensor could detect the target bacteria with a detection limit of 4×10³CFU/mL. A linear response was found between 4×10³- 4×10⁶CFU/mL. This biosensor was characterized with high sensitivity, excellent selectivity, short detection time and easy operation It has a promising application in clinical laboratory diagnoses, environmental detection and food safety.

Detection of Prosthetic Hip Infection at Revision Arthroplasty by Immunofluorescence Microscopy and PCR Amplification of the Bacterial 16S rRNA Gene

PubMed Central

Tunney, Michael M.; Patrick, Sheila; Curran, Martin D.; Ramage, Gordon; Hanna, Donna; Nixon, James R.; Gorman, Sean P.; Davis, Richard I.; Anderson, Neil

1999-01-01

In this study the detection rates of bacterial infection of hip prostheses by culture and nonculture methods were compared for 120 patients with total hip revision surgery. By use of strict anaerobic bacteriological practice during the processing of samples and without enrichment, the incidence of infection by culture of material dislodged from retrieved prostheses after ultrasonication (sonicate) was 22%. Bacteria were observed by immunofluorescence microscopy in 63% of sonicate samples with a monoclonal antibody specific for Propionibacterium acnes and polyclonal antiserum specific for Staphylococcus spp. The bacteria were present either as single cells or in aggregates of up to 300 bacterial cells. These aggregates were not observed without sonication to dislodge the biofilm. Bacteria were observed in all of the culture-positive samples, and in some cases in which only one type of bacterium was identified by culture, both coccoid and coryneform bacteria were observed by immunofluorescence microscopy. Bacteria from skin-flake contamination were readily distinguishable from infecting bacteria by immunofluorescence microscopy. Examination of skin scrapings did not reveal large aggregates of bacteria but did reveal skin cells. These were not observed in the sonicates. Bacterial DNA was detected in 72% of sonicate samples by PCR amplification of a region of the bacterial 16S rRNA gene with universal primers. All of the culture-positive samples were also positive for bacterial DNA. Evidence of high-level infiltration either of neutrophils or of lymphocytes or macrophages into associated tissue was observed in 73% of patients. Our results indicate that the incidence of prosthetic joint infection is grossly underestimated by current culture detection methods. It is therefore imperative that current clinical practice with regard to the detection and subsequent treatment of prosthetic joint infection be reassessed in the light of these results. PMID:10488193

The use of epifluorescent microscopy and quantitative polymerase chain reaction to determine the presence/absence and identification of microorganisms associated with domestic and foreign wallboard samples

USGS Publications Warehouse

Griffin, Dale W.

2011-01-01

Epifluorescent microscopy and quantitative polymerase chain reaction (qPCR) were utilized to determine the presence, concentration and identification of bacteria, and more specifically sulfate reducing bacteria (SRB) in subsamples of Chinese and North American wallboard, and wallboard-mine rock. Bacteria were visible in most subsamples, which included wallboard-lining paper from each side of the wallboard, wallboard filler, wallboard tape and fragments of mined wallboard rock via microscopy. Observed bacteria occurred as single or small clusters of cells and no mass aggregates indicating colonization were noted. Universal 16S qPCR was utilized to directly examine samples and detected bacteria at concentrations ranging from 1.4 x 103 to 6.4 x 104 genomic equivalents per mm2 of paper or per gram of wallboard filler or mined rock, in 12 of 41 subsamples. Subsamples were incubated in sulfate reducing broth for ~30 to 60 days (enrichment assay) and then analyzed by universal 16S and SRB qPCR. Enrichment universal 16S qPCR detected bacteria in 32 of 41 subsamples at concentrations ranging from 1.5 x 104 to 4.2 x 107 genomic equivalents per ml of culture broth. Evaluation of enriched subsamples by SRB qPCR demonstrated that SRB were not detectable in most of the samples and if they were detected, detection was not reproducible (an indication of low concentrations, if present). Enrichment universal 16S and SRB qPCR demonstrated that viable bacteria were present in subsamples (as expected given exposure of the samples following manufacture, transport and use) but that SRB were either not present or present at very low numbers. Further, no differences in trends were noted between the various Chinese and North American wallboard samples. In all, the microscopy and qPCR data indicated that the suspected ‘sulfur emissions’ emanating from suspect wallboard samples is not due to microbial activity.

Time Resolved Microfluorescence In Biomedical Diagnosis

NASA Astrophysics Data System (ADS)

Schneckenburger, Herbert

1985-12-01

A measuring system combining subnanosecond laser-induced fluorescence with microscopic signal detection was installed and used for diverse projects in the biomedical and environmental fields. These projects range from tumor diagnosis and enzymatic analysis to measurements of the activity of methanogenic bacteria, which affect biogas production and waste water cleaning. The advantages of this method and its practical applicability are discussed.

Nucleic acid extraction from polluted estuarine water for detection of viruses and bacteria by PCR and RT-PCR analysis.

PubMed

Petit, F; Craquelin, S; Guespin-Michel, J; Buffet-Janvresse, C

1999-03-01

We describe an extraction protocol for genomic DNA and RNA of both viruses and bacteria from polluted estuary water. This procedure was adapted to the molecular study of microflora of estuarine water where bacteria and viruses are found free, forming low-density biofilms, or intimately associated with organo-mineral particles. The sensitivity of the method was determined with seeded samples for RT-PCR and PCR analysis of viruses (10 virions/mL), and bacteria (1 colony-forming unit mL). We report an example of molecular detection of both poliovirus and Salmonella in the Seine estuary (France) and an approach to studying their association with organo-mineral particles.

Effectiveness of onsite wastewater reuse system in reducing bacterial contaminants measured with human-specific IMS/ATP and qPCR.

PubMed

Agidi, Senyo; Vedachalam, Sridhar; Mancl, Karen; Lee, Jiyoung

2013-01-30

Water shortages and the drive to recycle is increasing interest in reuse of reclaimed wastewater. Timely and cost-effective ways to detect fecal pollutants prior to reuse increases confidence of residents and neighbors concerned about reuse of reclaimed wastewater. The on-site wastewater treatment and reuse systems (OWTRS) used in this study include a septic tank, peat bioreactor, ClO(2) disinfection and land spray irrigation system. Bacteroides fragilis, Escherichia coli and Enterococcus spp., were tested with immunomagnetic separation/ATP bioluminescence (IMS/ATP), qPCR and culture-based methods. The results displayed a 2-log reduction in fecal bacteria in the peat bioreactor and a 5-log reduction following chloride dioxide disinfection. The fecal bacteria levels measured by IMS/ATP correlated with qPCR results: HuBac 16S (R(2) = 0.903), Bf-group 16S (R(2) = 0.956), gyrB (R(2) = 0.673), and Ent 23S (R(2) = 0.724). This is the first study in which the newly developed human-specific IMS/ATP and previously developed IMS/ATP were applied for determining OWTRS efficiency. Results of the study revealed that IMS/ATP is a timely and cost-effective way to detect fecal contaminants, and results were validated with qPCR and culture based methods. The new IMS/ATP can also be applied broadly in the detection of human-originated fecal contamination. Copyright © 2012 Elsevier Ltd. All rights reserved.

Rapid identification of antibiotic resistance using droplet microfluidics.

PubMed

Keays, Marie C; O'Brien, Mark; Hussain, Anam; Kiely, Patrick A; Dalton, Tara

2016-04-02

Culturing bacteria and monitoring bacterial cell growth is a critical issue when dealing with patients who present with bacterial infections. One of the main challenges that arises is the time taken to identify the particular strain of bacteria and consequently, decide the correct treatment. In the majority of cases, broad spectrum antibiotics are used to target infections when a narrow spectrum drug would be more appropriate. The efficient monitoring of bacterial growth and potential antibiotic resistance is necessary to identify the best treatment options for patients. Minturising the reactions into microfluidic droplets offers a novel method to rapidy analyze bacteria. Microfluidics facilitates low volume reactions that provide a unique system where each droplet reaction acts as an individual bioreactor. Here, we designed and built a novel platform that allowed us to create and monitor E.coli microfluidic droplet cultures. Optical capacity was built in and measurements of bacterial cultures were captured facilitating the continuous monitoring of individual reactions. The capacity of the instrument was demonstrated by the application of treatments to both bacteria and drug resistant strains of bacteria. We were able to detect responses within one hour in the droplet cultures, demonstrating the capacity of this workflow to the culture and rapid characterization of bacterial strains.

Long-term effect of pH on short-chain fatty acids accumulation and microbial community in sludge fermentation systems.

PubMed

Yuan, Yue; Wang, Shuying; Liu, Ye; Li, Baikun; Wang, Bo; Peng, Yongzhen

2015-12-01

Long-term effect of pH (4, 10, and uncontrolled) on short-chain fatty acid (SCFA) accumulation, microbial community and sludge reduction were investigated in waste activated sludge (WAS) fermentors for over 90days. The average SCFAs accumulation was 1721.4 (at pH 10), 114.2 (at pH 4), and 58.1 (at uncontrolled pH) mg chemical oxygen demand (COD)/L. About 31.65mgCOD/L was produced at pH 10, accounting for 20% of the influent COD. Illumina MiSeq sequencing revealed that Alcaligenes (hydrolic bacteria) and Erysipelothrix (acidogenic bacteria) were enriched at pH 10, while less acidogenic bacteria existed at pH 4 than pH 10, and no acidogenic bacteria were detected at the uncontrolled pH. The ratios of archaea to bacteria were 1:41, 1:16, and 1:9 at the pH of 10, 4, and uncontrolled. This study elucidated the effects of pH on WAS fermentation, and established the correlation of microbial structure with SCFAs accumulations and sludge reduction. Copyright © 2015 Elsevier Ltd. All rights reserved.

Cross-contamination potential of saliva ejectors used in dentistry.

PubMed

Barbeau, J; ten Bokum, L; Gauthier, C; Prévost, A P

1998-12-01

It has been postulated that evacuation systems used in dentistry could be a source of cross-contamination between patients through backflow of bacteria dislodged from the saliva ejector tubings. The bacterial microflora associated with these systems was characterized using transmission electron microscopy (TEM) and microbiological cultures. The potential for backflow was investigated by a study of pressure differentials in evacuation system tubing and by the presence of bacteria in backflow samples. Evacuation lines were coated with microbial biofilms in which microcolonies of Gram-positive cocci and Gram-negative bacilli predominated, embedded in an extensive polysaccharide matrix. Most bacteria were metabolically active. Occasionally, buccal material such as collagen, fibrin and eukaryotic cell debris was observed. In other experiments, flow reversal was detected several times during saliva ejector use though each of these events was brief (less than 0.1 s). Aspiration of saliva, or occlusion of the mouthpiece opening by the oral mucosa, were the major factors leading to backflow episodes. Bacteria associated with backflow were found in almost 25% assays, with counts ranging from 1-300 cfu/occurrence. The majority of the bacteria isolated from biofilm or backflow samples were staphylococci, micrococci and non-fermentive Gram-negative rods. Pathogens such as Pseudomonas aeruginosa and Staphylococcus aureus were also isolated from backflow fluids. No oral streptococci could be recovered from biofilms in the tubing beyond 15 min from the last saliva ejector use however, suggesting that these species did not survive in the biofilms. These data suggest, although without direct proof of cross-contamination, the possible existence of an infectious risk associated with oral evacuation systems, as potential pathogens may be shed from tubing biofilms following backflow. Even if the risk of cross-contamination between patients is considered to be low, the necessity for regular disinfection of these systems must be stressed, since biofilms can serve as a reservoir for pathogens or harbor potentially infectious material.

Clinical antibacterial effectiveness of the self-adjusting file system.

PubMed

Neves, M A S; Rôças, I N; Siqueira, J F

2014-04-01

To evaluate in vivo the antibacterial effectiveness of the self-adjusting file (SAF) using molecular methods. Root canals from single-rooted teeth with apical periodontitis were instrumented using the SAF system under continuous irrigation with 2.5% NaOCl. DNA extracts from samples taken before and after instrumentation were subjected to quantitative analysis of total bacteria counts and levels of streptococci by quantitative real-time polymerase chain reaction (qPCR). The reverse-capture checkerboard assay was also used to identify 28 bacterial taxa before (S1) and after (S2) SAF instrumentation. SAF was also compared with a conventional hand nickel-titanium instrumentation technique for total bacterial reduction. Data from qPCR were analysed statistically within groups using the Wilcoxon matched pairs test and between groups using the Mann-Whitney U-test and the Fisher's exact test, with significance level set at P < 0.05. Self-adjusting file significantly reduced the total bacterial counts from a mean number of 1.96 × 10(7) cells to 1.34 × 10(4) cells (P < 0.001). Quantitatively, the 99.9% reduction in total bacterial counts associated with the SAF system was significantly superior to the 95.1% reduction obtained by hand instrumentation (P < 0.001). Qualitatively, SAF resulted in significantly more cases with negative PCR results for bacteria (54.5%) than hand instrumentation (4.5%) (P < 0.001). The SAF system succeeded in significantly reducing the streptococcal levels, but four cases still harboured these bacteria in S2. Checkerboard analysis revealed that not only streptococci but also some anaerobic and even as-yet-uncultivated bacteria may resist the effects of chemomechanical procedures. The SAF instrumentation system was highly effective in reducing bacterial populations from infected root canals and performed significantly better than hand instrumentation. However, because half of the samples still had detectable bacteria after preparation with SAF, supplementary disinfection is still required to maximize bacterial elimination. © 2013 International Endodontic Journal. Published by John Wiley & Sons Ltd.

Simple, rapid, and cost-effective modified Carba NP test for carbapenemase detection among Gram-negative bacteria

PubMed Central

Rudresh, Shoorashetty Manohara; Ravi, Giriyapur Siddappa; Sunitha, Lakshminarayanappa; Hajira, Sadiya Noor; Kalaiarasan, Ellappan; Harish, Belgode Narasimha

2017-01-01

PURPOSE: Detection of carbapenemases among Gram-negative bacteria (GNB) is important for both clinicians and infection control practitioners. The Clinical and Laboratory Standards Institute recommends Carba NP (CNP) as confirmatory test for carbapenemase production. The reagents required for CNP test are costly and hence the test cannot be performed on a routine basis. The present study evaluates modifications of CNP test for rapid detection of carbapenemases among GNB. MATERIALS AND METHODS: The GNB were screened for carbapenemase production using CNP, CarbAcineto NP (CANP), and modified CNP (mCNP) test. A multiplex polymerase chain reaction (PCR) was performed on all the carbapenem-resistant bacteria for carbapenemase genes. The results of three phenotypic tests were compared with PCR. RESULTS: A total of 765 gram negative bacteria were screened for carbapenem resistance. Carbapenem resistance was found in 144 GNB. The metallo-β-lactamases were most common carbapenemases followed by OXA-48-like enzymes. The CANP test was most sensitive (80.6%) for carbapenemases detection. The mCNP test was 62.1% sensitive for detection of carbapenemases. The mCNP, CNP, and CANP tests were equally sensitive (95%) for detection of NDM enzymes among Enterobacteriaceae. The mCNP test had poor sensitivity for detection of OXA-48-like enzymes. CONCLUSION: The mCNP test was rapid, cost-effective, and easily adoptable on routine basis. The early detection of carbapenemases using mCNP test will help in preventing the spread of multidrug-resistant organisms in the hospital settings. PMID:28966495

Simple, rapid, and cost-effective modified Carba NP test for carbapenemase detection among Gram-negative bacteria.

PubMed

Rudresh, Shoorashetty Manohara; Ravi, Giriyapur Siddappa; Sunitha, Lakshminarayanappa; Hajira, Sadiya Noor; Kalaiarasan, Ellappan; Harish, Belgode Narasimha

2017-01-01

Detection of carbapenemases among Gram-negative bacteria (GNB) is important for both clinicians and infection control practitioners. The Clinical and Laboratory Standards Institute recommends Carba NP (CNP) as confirmatory test for carbapenemase production. The reagents required for CNP test are costly and hence the test cannot be performed on a routine basis. The present study evaluates modifications of CNP test for rapid detection of carbapenemases among GNB. The GNB were screened for carbapenemase production using CNP, CarbAcineto NP (CANP), and modified CNP (mCNP) test. A multiplex polymerase chain reaction (PCR) was performed on all the carbapenem-resistant bacteria for carbapenemase genes. The results of three phenotypic tests were compared with PCR. A total of 765 gram negative bacteria were screened for carbapenem resistance. Carbapenem resistance was found in 144 GNB. The metallo-β-lactamases were most common carbapenemases followed by OXA-48-like enzymes. The CANP test was most sensitive (80.6%) for carbapenemases detection. The mCNP test was 62.1% sensitive for detection of carbapenemases. The mCNP, CNP, and CANP tests were equally sensitive (95%) for detection of NDM enzymes among Enterobacteriaceae. The mCNP test had poor sensitivity for detection of OXA-48-like enzymes. The mCNP test was rapid, cost-effective, and easily adoptable on routine basis. The early detection of carbapenemases using mCNP test will help in preventing the spread of multidrug-resistant organisms in the hospital settings.

Impact of Manure Fertilization on the Abundance of Antibiotic-Resistant Bacteria and Frequency of Detection of Antibiotic Resistance Genes in Soil and on Vegetables at Harvest

PubMed Central

Marti, Romain; Scott, Andrew; Tien, Yuan-Ching; Murray, Roger; Sabourin, Lyne; Zhang, Yun

2013-01-01

Consumption of vegetables represents a route of direct human exposure to bacteria found in soil. The present study evaluated the complement of bacteria resistant to various antibiotics on vegetables often eaten raw (tomato, cucumber, pepper, carrot, radish, lettuce) and how this might vary with growth in soil fertilized inorganically or with dairy or swine manure. Vegetables were sown into field plots immediately following fertilization and harvested when of marketable quality. Vegetable and soil samples were evaluated for viable antibiotic-resistant bacteria by plate count on Chromocult medium supplemented with antibiotics at clinical breakpoint concentrations. DNA was extracted from soil and vegetables and evaluated by PCR for the presence of 46 gene targets associated with plasmid incompatibility groups, integrons, or antibiotic resistance genes. Soil receiving manure was enriched in antibiotic-resistant bacteria and various antibiotic resistance determinants. There was no coherent corresponding increase in the abundance of antibiotic-resistant bacteria enumerated from any vegetable grown in manure-fertilized soil. Numerous antibiotic resistance determinants were detected in DNA extracted from vegetables grown in unmanured soil. A smaller number of determinants were additionally detected on vegetables grown only in manured and not in unmanured soil. Overall, consumption of raw vegetables represents a route of human exposure to antibiotic-resistant bacteria and resistance determinants naturally present in soil. However, the detection of some determinants on vegetables grown only in freshly manured soil reinforces the advisability of pretreating manure through composting or other stabilization processes or mandating offset times between manuring and harvesting vegetables for human consumption. PMID:23851089

Quantification of Propionic Acid in the Bovine Spinal Disk After Infection of the Tissue With Propionibacteria acnes Bacteria.

PubMed

Magnitsky, Sergey; Dudli, Stefan; Tang, Xinyan; Kaur, Jaskanwaljeet; Diaz, Joycelyn; Miller, Steve; Lotz, Jeffrey C

2018-06-01

Research. The goal of this study was to investigate whether Propionibacteria acnes infection of the intervertebral disc can be detected noninvasively by nuclear magnetic resonance (NMR) spectroscopy. Microbiological studies of surgical samples suggest that a significant subpopulation of back pain patients may have occult disc infection with P. acnes bacteria. This hypothesis is further supported by a double-blind clinical trial showing that back pain patients with Modic type 1 changes may respond to antibiotic treatment. Because significant side effects are associated with antibiotic treatment, there is a need for a noninvasive method to detect whether specific discs in back pain patients are infected with P acnes bacteria. P. acnes bacteria were obtained from human patients. NMR detection of a propionic acid (PA) in the bacteria extracts was conducted on 500 MHz high-resolution spectrometer, whereas in vivo NMR spectroscopy of an isolated bovine disk tissue infected with P. acnes was conducted on 7 T magnetic resonance imaging scanner. NMR spectra of P. acnes metabolites revealed a distinct NMR signal with identical chemical shits (1.05 and 2.18 ppm) as PA (a primary P. acne metabolite). The 1.05 ppm signal does not overlap with other bacteria metabolites, and its intensity increases linearly with P. acnes concentration. Bovine disks injected with P. acnes bacteria revealed a very distinct NMR signal at 1.05 ppm, which linearly increased with P. acnes concentration. The 1.05 ppm NMR signal from PA can be used as a marker of P. acnes infection of discs. This signal does not overlap with other disc metabolites and linearly depends on P. acnes concentration. Consequently, NMR spectroscopy may provide a noninvasive method to detect disc infection in the clinical setting. N/A.

Polydiacetylene-Based Liposomes: An "Optical Tongue" for Bacteria Detection and Identification

ERIC Educational Resources Information Center

West, Matthew R.; Hanks, Timothy W.; Watson, Rhett T.

2009-01-01

Food- and water-borne bacteria are a major health concern worldwide. Current detection methods are time-consuming and require sophisticated equipment that is not always readily available. However, new techniques based on nanotechnology are under development that will result in a new generation of sensors. In this experiment, liposomes are…

Application of leftover sample material from waterborne protozoa monitoring for the molecular detection of Bacteroidales and fecal source tracking markers

EPA Science Inventory

In this study, we examined the potential for detecting fecal bacteria and microbial source tracking markers in samples discarded during the concentration of Cryptosporidium and Giardia using USEPA Method 1623. Recovery rates for different fecal bacteria were determined using sp...

Impact of Strain Variation on the Ability of Biosensor Technology to Detect Salmonella enterica

USDA-ARS?s Scientific Manuscript database

Introduction: It is important to develop methods that can quickly and accurately detect the presence of bacteria in the food supply that cause disease. Salmonella enterica is a bacteria that is often associated with contamination of food. Strains vary in their ability to cause illness and to spread...

Detecting bacteria in food- harder than searching for a needle in a haystack

USDA-ARS?s Scientific Manuscript database

Actually, I think it would have been easier to find a needle in a haystack than to locate variables pertaining to that time-honored question and mathematically compute the relative difficulty of detecting bacteria versus searching for aforementioned needle. Perhaps it might be unusual given that I ...

Fast detection of Listeria monocytogenes through a nanohybrid quantum dot complex.

PubMed

Donoso, Wendy; Castro, Ricardo I; Guzmán, Luis; López-Cabaña, Zoraya; Nachtigall, Fabiane M; Santos, Leonardo S

2017-09-01

Listeria monocytogenes is a recognized foodborne pathogen that causes listeriosis in susceptible consumers. Currently, the detection systems for Listeria in food detect live and dead bacteria, being the viable microorganisms most relevant for their ability to cause sickness in the population at risk. For this reason, a new nanohybrid compound was developed for the optical detection of Listeria that was based on polyamidoamine dendrimers functionalized with an auxotrophic cofactor (lipoic acid), together with the coupling of fluorescent semiconductor crystals (quantum dots). The nanohybrid sensor has a detection limit for viable L. monocytogenes of 5.19 × 10 3 colony-forming units per milliliter under epifluorescence microscopy. It was specific when used among other pathogens commonly found in food.

Atrazine, chlorpyrifos, and iprodione effect on the biodiversity of bacteria, actinomycetes, and fungi in a pilot biopurification system with a green cover.

PubMed

Elgueta, Sebastian; Correa, Arturo; Campo, Marco; Gallardo, Felipe; Karpouzas, Dimitrios; Diez, Maria Cristina

2017-09-02

The use of biopurification systems can mitigate the effects of pesticide contamination on farms. The primary aim of this study was to evaluate the effect of pesticide dissipation on microbial communities in a pilot biopurification system. The pesticide dissipation of atrazine, chlorpyrifos and iprodione (35 mg kg -1 active ingredient [a.i.]) and biological activity were determined for 40 days. The microbial communities (bacteria, actinomycetes and fungi) were analyzed using denaturing gradient gel electrophoresis (DGGE). In general, pesticide dissipation was the highest by day 5 and reached 95%. The pesticides did not affect biological activity during the experiment. The structure of the actinomycete and bacterial communities in the rhizosphere was more stable during the evaluation than that in the communities in the control without pesticides. The rhizosphere fungal communities, detected using DGGE, showed small and transitory shifts with time. To conclude, rhizosphere microbial communities were not affected during pesticide dissipation in a pilot biopurification system.

Streptavidin mutants

DOEpatents

Sano, Takeshi; Cantor, Charles R.; Vajda, Sandor; Reznik, Gabriel O.; Smith, Cassandra L.; Pandori, Mark W.

2000-01-01

The present invention relates to streptavidin proteins and peptides having a altered physical properties such as an increased stability or increased or decreased affinity for binding biotin. The invention also relates to methods for the detection, identification, separation and isolation of targets using streptavidin proteins or peptides. Streptavidin with increased or reduced affinity allows for the use of the streptavidin-biotin coupling systems for detection and isolation systems wherein it is necessary to remove of one or the other of the binding partners. Such systems are useful for the purification of functional proteins and viable cells. The invention also relates to nucleic acids which encode these streptavidin proteins and peptides and to recombinant cells such as bacteria, yeast and mammalian cells which contain these nucleic acids.

Next-generation sequencing identification of pathogenic bacterial genes and their relationship with fecal indicator bacteria in different water sources in the Kathmandu Valley, Nepal.

PubMed

Ghaju Shrestha, Rajani; Tanaka, Yasuhiro; Malla, Bikash; Bhandari, Dinesh; Tandukar, Sarmila; Inoue, Daisuke; Sei, Kazunari; Sherchand, Jeevan B; Haramoto, Eiji

2017-12-01

Bacteriological analysis of drinking water leads to detection of only conventional fecal indicator bacteria. This study aimed to explore and characterize bacterial diversity, to understand the extent of pathogenic bacterial contamination, and to examine the relationship between pathogenic bacteria and fecal indicator bacteria in different water sources in the Kathmandu Valley, Nepal. Sixteen water samples were collected from shallow dug wells (n=12), a deep tube well (n=1), a spring (n=1), and rivers (n=2) in September 2014 for 16S rRNA gene next-generation sequencing. A total of 525 genera were identified, of which 81 genera were classified as possible pathogenic bacteria. Acinetobacter, Arcobacter, and Clostridium were detected with a relatively higher abundance (>0.1% of total bacterial genes) in 16, 13, and 5 of the 16 samples, respectively, and the highest abundance ratio of Acinetobacter (85.14%) was obtained in the deep tube well sample. Furthermore, the bla OXA23-like genes of Acinetobacter were detected using SYBR Green-based quantitative PCR in 13 (35%) of 37 water samples, including the 16 samples that were analyzed for next-generation sequencing, with concentrations ranging 5.3-7.5logcopies/100mL. There was no sufficient correlation found between fecal indicator bacteria, such as Escherichia coli and total coliforms, and potential pathogenic bacteria, as well as the bla OXA23-like gene of Acinetobacter. These results suggest the limitation of using conventional fecal indicator bacteria in evaluating the pathogenic bacteria contamination of different water sources in the Kathmandu Valley. Copyright © 2017 Elsevier B.V. All rights reserved.

Molecular methods routinely used to detect Coxiella burnetii in ticks cross-react with Coxiella-like bacteria

PubMed Central

Elsa, Jourdain; Duron, Olivier; Séverine, Barry; González-Acuña, Daniel; Sidi-Boumedine, Karim

2015-01-01

Background Q fever is a widespread zoonotic disease caused by Coxiella burnetii. Ticks may act as vectors, and many epidemiological studies aim to assess C. burnetii prevalence in ticks. Because ticks may also be infected with Coxiella-like bacteria, screening tools that differentiate between C. burnetii and Coxiella-like bacteria are essential. Methods In this study, we screened tick specimens from 10 species (Ornithodoros rostratus, O. peruvianus, O. capensis, Ixodes ricinus, Rhipicephalus annulatus, R. decoloratus, R. geigy, O. sonrai, O. occidentalis, and Amblyomma cajennense) known to harbor specific Coxiella-like bacteria, by using quantitative PCR primers usually considered to be specific for C. burnetii and targeting, respectively, the IS1111, icd, scvA, p1, and GroEL/htpB genes. Results We found that some Coxiella-like bacteria, belonging to clades A and C, yield positive PCR results when screened with primers initially believed to be C. burnetii-specific. Conclusions These results suggest that PCR-based surveys that aim to detect C. burnetii in ticks by using currently available methods must be interpreted with caution if the amplified products cannot be sequenced. Future molecular methods that aim at detecting C. burnetii need to take into account the possibility that cross-reactions may exist with Coxiella-like bacteria. PMID:26609691

Molecular methods routinely used to detect Coxiella burnetii in ticks cross-react with Coxiella-like bacteria.

PubMed

Elsa, Jourdain; Duron, Olivier; Séverine, Barry; González-Acuña, Daniel; Sidi-Boumedine, Karim

2015-01-01

Q fever is a widespread zoonotic disease caused by Coxiella burnetii. Ticks may act as vectors, and many epidemiological studies aim to assess C. burnetii prevalence in ticks. Because ticks may also be infected with Coxiella-like bacteria, screening tools that differentiate between C. burnetii and Coxiella-like bacteria are essential. In this study, we screened tick specimens from 10 species (Ornithodoros rostratus, O. peruvianus, O. capensis, Ixodes ricinus, Rhipicephalus annulatus, R. decoloratus, R. geigy, O. sonrai, O. occidentalis, and Amblyomma cajennense) known to harbor specific Coxiella-like bacteria, by using quantitative PCR primers usually considered to be specific for C. burnetii and targeting, respectively, the IS1111, icd, scvA, p1, and GroEL/htpB genes. We found that some Coxiella-like bacteria, belonging to clades A and C, yield positive PCR results when screened with primers initially believed to be C. burnetii-specific. These results suggest that PCR-based surveys that aim to detect C. burnetii in ticks by using currently available methods must be interpreted with caution if the amplified products cannot be sequenced. Future molecular methods that aim at detecting C. burnetii need to take into account the possibility that cross-reactions may exist with Coxiella-like bacteria.

Enteric Viruses and Fecal Bacteria Indicators to Assess Groundwater Quality and Suitability for Irrigation

PubMed Central

De Giglio, Osvalda; Caggiano, Giuseppina; Bagordo, Francesco; Barbuti, Giovanna; Brigida, Silvia; Lugoli, Federica; Grassi, Tiziana; La Rosa, Giuseppina; Lucentini, Luca; Uricchio, Vito Felice; De Donno, Antonella; Montagna, Maria Teresa

2017-01-01

According to Italian Ministerial Decree No. 185 of 12 June 2003, water is considered suitable for irrigation if levels of fecal bacteria (i.e., Escherichia coli and Salmonella) are within certain parameters. The detection of other microorganisms is not required. The aim of this study is to determine the bacteriological quality of groundwater used for irrigation and the occurrence of enteric viruses (Norovirus, Enterovirus, Rotavirus, Hepatovirus A), and to compare the presence of viruses with the fecal bacteria indicators. A total of 182 wells was analyzed. Widespread fecal contamination of Apulian aquifers was detected (141 wells; 77.5%) by the presence of fecal bacteria (i.e., E. coli, Salmonella, total coliforms, and enterococci). Considering bacteria included in Ministerial Decree No. 185, the water from 35 (19.2%) wells was unsuitable for irrigation purposes. Among 147 wells with water considered suitable, Norovirus, Rotavirus, and Enterovirus were detected in 23 (15.6%) wells. No Hepatovirus A was isolated. Consequently, 58 wells (31.9%) posed a potential infectious risk for irrigation use. This study revealed the inadequacy of fecal bacteria indicators to predict the occurrence of viruses in groundwater and it is the first in Italy to describe the presence of human rotaviruses in well water used for irrigation. PMID:28538682

Cationic polyelectrolyte functionalized magnetic particles assisted highly sensitive pathogens detection in combination with polymerase chain reaction and capillary electrophoresis.

PubMed

Chen, Jia; Lin, Yuexin; Wang, Yu; Jia, Li

2015-06-01

Pathogenic bacteria cause significant morbidity and mortality to humans. There is a pressing need to establish a simple and reliable method to detect them. Herein, we show that magnetic particles (MPs) can be functionalized by poly(diallyl dimethylammonium chloride) (PDDA), and the particles (PDDA-MPs) can be utilized as adsorbents for capture of pathogenic bacteria from aqueous solution based on electrostatic interaction. The as-prepared PDDA-MPs were characterized by Fourier-transform infrared spectroscopy, zeta potential, vibrating sample magnetometry, X-ray diffraction spectrometry, scanning electron microscopy, and transmission electron microscopy. The adsorption equilibrium time can be achieved in 3min. According to the Langmuir adsorption isotherm, the maximum adsorption capacities for E. coli O157:H7 (Gram-negative bacteria) and L. monocytogenes (Gram-positive bacteria) were calculated to be 1.8×10(9) and 3.1×10(9)cfumg(-1), respectively. The bacteria in spiked mineral water (1000mL) can be completely captured when applying 50mg of PDDA-MPs and an adsorption time of 5min. In addition, PDDA-MPs-based magnetic separation method in combination with polymerase chain reaction and capillary electrophoresis allows for rapid detection of 10(1)cfumL(-1) bacteria. Copyright © 2015 Elsevier B.V. All rights reserved.

Bacteria and bacterial DNA in atherosclerotic plaque and aneurysmal wall biopsies from patients with and without periodontitis

PubMed Central

Armingohar, Zahra; Jørgensen, Jørgen J.; Kristoffersen, Anne Karin; Abesha-Belay, Emnet; Olsen, Ingar

2014-01-01

Background Several studies have reported an association between chronic periodontitis (CP) and cardiovascular diseases. Detection of periodontopathogens, including red complex bacteria (RCB), in vascular lesions has suggested these bacteria to be involved in the pathogenesis of atherosclerosis and abdominal aortic aneurysms. Objective In this study, we investigate bacteria and their DNA in vascular biopsies from patients with vascular diseases (VD; i.e. abdominal aortic aneurysms, atherosclerotic carotid, and common femoral arteries), with and without CP. Methods DNA was extracted from vascular biopsies selected from 40 VD patients: 30 with CP and 10 without CP. The V3-V5 region of the 16S rDNA (V3-V5) was polymerase chain reaction (PCR)-amplified, and the amplicons were cloned into Escherichia coli, sequenced, and classified (GenBank and the Human Oral Microbiome database). Species-specific primers were used for the detection of Porphyromonas gingivalis. In addition, 10 randomly selected vascular biopsies from the CP group were subjected to scanning electron microscopy (SEM) for visualization of bacteria. Checkerboard DNA–DNA hybridization was performed to assess the presence of RCB in 10 randomly selected subgingival plaque samples from CP patients. Results A higher load and mean diversity of bacteria were detected in vascular biopsies from VD patients with CP compared to those without CP. Enterobacteriaceae were frequently detected in vascular biopsies together with cultivable, commensal oral, and not-yet-cultured bacterial species. While 70% of the subgingival plaque samples from CP patients showed presence of RCB, only P. gingivalis was detected in one vascular biopsy. Bacterial cells were seen in all 10 vascular biopsies examined by SEM. Conclusions A higher bacterial load and more diverse colonization were detected in VD lesions of CP patients as compared to patients without CP. This indicated that a multitude of bacterial species both from the gut and the oral cavity, rather than exclusively periodontopathogens, may be involved as additional risk factors in the pathogenesis of VD. PMID:25006361

Bacteria-instructed synthesis of polymers for self-selective microbial binding and labelling

PubMed Central

Magennis, E. Peter; Fernandez-Trillo, Francisco; Sui, Cheng; Spain, Sebastian G.; Bradshaw, David; Churchley, David; Mantovani, Giuseppe; Winzer, Klaus; Alexander, Cameron

2014-01-01

The detection and inactivation of pathogenic strains of bacteria continues to be an important therapeutic goal. Hence, there is a need for materials that can bind selectively to specific microorganisms, for diagnostic or anti-infective applications, but which can be formed from simple and inexpensive building blocks. Here, we exploit bacterial redox systems to induce a copper-mediated radical polymerisation of synthetic monomers at cell surfaces, generating polymers in situ that bind strongly to the microorganisms which produced them. This ‘bacteria-instructed synthesis’ can be carried out with a variety of microbial strains, and we show that the polymers produced are self-selective binding agents for the ‘instructing’ cell types. We further expand on the bacterial redox chemistries to ‘click’ fluorescent reporters onto polymers directly at the surfaces of a range of clinical isolate strains, allowing rapid, facile and simultaneous binding and visualisation of pathogens. PMID:24813421

Bacteria-instructed synthesis of polymers for self-selective microbial binding and labelling

NASA Astrophysics Data System (ADS)

Magennis, E. Peter; Fernandez-Trillo, Francisco; Sui, Cheng; Spain, Sebastian G.; Bradshaw, David J.; Churchley, David; Mantovani, Giuseppe; Winzer, Klaus; Alexander, Cameron

2014-07-01

The detection and inactivation of pathogenic strains of bacteria continues to be an important therapeutic goal. Hence, there is a need for materials that can bind selectively to specific microorganisms for diagnostic or anti-infective applications, but that can be formed from simple and inexpensive building blocks. Here, we exploit bacterial redox systems to induce a copper-mediated radical polymerization of synthetic monomers at cell surfaces, generating polymers in situ that bind strongly to the microorganisms that produced them. This ‘bacteria-instructed synthesis’ can be carried out with a variety of microbial strains, and we show that the polymers produced are self-selective binding agents for the ‘instructing’ cell types. We further expand on the bacterial redox chemistries to ‘click’ fluorescent reporters onto polymers directly at the surfaces of a range of clinical isolate strains, allowing rapid, facile and simultaneous binding and visualization of pathogens.

Infection of Immature Ixodes scapularis (Acari: Ixodidae) by Membrane Feeding

PubMed Central

Oliver, Jonathan D.; Lynn, Geoffrey E.; Burkhardt, Nicole Y.; Price, Lisa D.; Nelson, Curtis M.; Kurtti, Timothy J.; Munderloh, Ulrike G.

2016-01-01

Abstract A reduction in the use of animals in infectious disease research is desirable for animal welfare as well as for simplification and standardization of experiments. An artificial silicone-based membrane-feeding system was adapted for complete engorgement of adult and nymphal Ixodes scapularis Say (Acari: Ixodidae), and for infecting nymphs with pathogenic, tick-borne bacteria. Six wild-type and genetically transformed strains of four species of bacteria were inoculated into sterile bovine blood and fed to ticks. Pathogens were consistently detected in replete nymphs by polymerase chain reaction. Adult ticks that ingested bacteria as nymphs were evaluated for transstadial transmission. Borrelia burgdorferi and Ehrlichia muris -like agent showed high rates of transstadial transmission to adult ticks, whereas Anaplasma phagocytophilum and Rickettsia monacensis demonstrated low rates of transstadial transmission/maintenance. Artificial membrane feeding can be used to routinely maintain nymphal and adult I. scapularis , and infect nymphs with tick-borne pathogens. PMID:26721866

Changes in Microbial Communities, Including both Uncultured and Culturable Bacteria, with Mid-Ocean Ballast-Water Exchange during a Voyage from Japan to Australia

PubMed Central

Tomaru, Akiko; Kawachi, Masanobu; Demura, Mikihide; Fukuyo, Yasuwo

2014-01-01

We assessed changes in the microbial communities in ballast water during a trans-Pacific voyage from Japan to Australia that included a mid-ocean ballast-water exchange. Uncultured (i.e., total) and culturable bacteria were counted and were characterized by using denaturing gradient gel electrophoresis (DGGE). There was a clear decrease over time in numbers of uncultured microorganisms, except for heterotrophic nanoflagellates, whereas the abundance of culturable bacteria initially decreased after the ballast-water exchange but then increased. The increase, however, was only up to 5.34% of the total number of uncultured bacteria. Cluster analysis showed that the DGGE profiles of uncultured bacteria clearly changed after the exchange. In contrast, there was no clear change in the DGGE profiles of culturable bacteria after the exchange. Multidimensional scaling analysis showed changes in microbial communities over the course of the voyage. Although indicator microbes as defined by the International Convention for the Control and Management of Ships' Ballast Water and Sediments were occasionally detected, no coliform bacteria were detected after the exchange. PMID:24817212

Environmental evaluation of coexistence of denitrifying anaerobic methane-oxidizing archaea and bacteria in a paddy field.

PubMed

Ding, Jing; Fu, Liang; Ding, Zhao-Wei; Lu, Yong-Ze; Cheng, Shuk H; Zeng, Raymond J

2016-01-01

The nitrate-dependent denitrifying anaerobic methane oxidation (DAMO) process, which is metabolized together by anaerobic methanotrophic archaea and NC10 phylum bacteria, is expected to be important for the global carbon and nitrogen cycles. However, there are little studies about the existence of this process and the functional microbes in environments. Therefore, the coexistence of DAMO archaea and bacteria in a paddy field was evaluated in this study. Next-generation sequencing showed that the two orders, Methanosarcinales and Nitrospirales, to which DAMO archaea and DAMO bacteria belong, were detected in the four soil samples. Then the in vitro experiments demonstrated both of nitrite- and nitrate-dependent DAMO activities, which confirmed the coexistence of DAMO archaea and DAMO bacteria. It was the first report about the coexistence of DAMO archaea and bacteria in a paddy field. Furthermore, anammox bacteria were detected in two of the four samples. The in vitro experiments did not show anammox activity in the initial period but showed low anammox activity after 20 days' enrichment. These results implicated that anammox bacteria may coexist with DAMO microorganisms in this field, but at a very low percentage.

Developing a Genetically Encoded, Cross-Species Biosensor for Detecting Ammonium and Regulating Biosynthesis of Cyanophycin.

PubMed

Xiao, Yi; Jiang, Wen; Zhang, Fuzhong

2017-10-20

Responding to nitrogen status is essential for all living organisms. Bacteria have evolved various complex and exquisite regulatory systems to control nitrogen metabolism. However, natural nitrogen regulatory systems, owing to their complexity, often function only in their original hosts and do not respond properly when transferred to another species. By harnessing the Lactococcus GlnRA system, we developed a genetically encoded, cross-species ammonium biosensor that displays a dynamic range up to 9-fold upon detection of ammonium ion. We demonstrated applications of this ammonium biosensor in three different species (Escherichia coli, Pseudomonas putida, and Synechocystis sp.) to detect different nitrogen sources. This ammonium sensor was further used to regulate the biosynthesis of a nitrogen-rich polymer, cyanophycin, based on ammonium concentration. Given the importance of nitrogen responses, the developed biosensor should be broadly applicable to synthetic biology and bioengineering.

Bovine Intestinal Bacteria Inactivate and Degrade Ceftiofur and Ceftriaxone with Multiple β-Lactamases▿

PubMed Central

Wagner, R. Doug; Johnson, Shemedia J.; Cerniglia, Carl E.; Erickson, Bruce D.

2011-01-01

The veterinary cephalosporin drug ceftiofur is rapidly degraded in the bovine intestinal tract. A cylinder-plate assay was used to detect microbiologically active ceftiofur, and high-performance liquid chromatography-mass spectrometry analysis was used to quantify the amount of ceftiofur remaining after incubation with bovine intestinal anaerobic bacteria, which were isolated from colon contents or feces from 8 cattle. Ninety-six percent of the isolates were able to inactivate ceftiofur to some degree, and 54% actually degraded the drug. None of 9 fungal isolates inactivated or degraded ceftiofur. Facultative and obligate anaerobic bacterial species that inactivated or degraded ceftiofur were identified with Vitek and Biolog systems, respectively. A subset of ceftiofur degraders also degraded the chemically similar drug ceftriaxone. Most of the species of bacteria that degraded ceftiofur belonged to the genera Bacillus and Bacteroides. PCR analysis of bacterial DNA detected specific β-lactamase genes. Bacillus cereus and B. mycoides isolates produced extended-spectrum β-lactamases and metallo-β-lactamases. Seven isolates of Bacteroides spp. produced multiple β-lactamases, including possibly CepA, and metallo-β-lactamases. Isolates of Eubacterium biforme, Bifidobacterium breve, and several Clostridium spp. also produced ceftiofur-degrading β-lactamases. An agar gel overlay technique on isoelectric focusing separations of bacterial lysates showed that β-lactamase enzymes were sufficient to degrade ceftiofur. These results suggest that ceftiofur is inactivated nonenzymatically and degraded enzymatically by multiple β-lactamases from bacteria in the large intestines of cattle. PMID:21876048

Bovine intestinal bacteria inactivate and degrade ceftiofur and ceftriaxone with multiple beta-lactamases.

PubMed

Wagner, R Doug; Johnson, Shemedia J; Cerniglia, Carl E; Erickson, Bruce D

2011-11-01

The veterinary cephalosporin drug ceftiofur is rapidly degraded in the bovine intestinal tract. A cylinder-plate assay was used to detect microbiologically active ceftiofur, and high-performance liquid chromatography-mass spectrometry analysis was used to quantify the amount of ceftiofur remaining after incubation with bovine intestinal anaerobic bacteria, which were isolated from colon contents or feces from 8 cattle. Ninety-six percent of the isolates were able to inactivate ceftiofur to some degree, and 54% actually degraded the drug. None of 9 fungal isolates inactivated or degraded ceftiofur. Facultative and obligate anaerobic bacterial species that inactivated or degraded ceftiofur were identified with Vitek and Biolog systems, respectively. A subset of ceftiofur degraders also degraded the chemically similar drug ceftriaxone. Most of the species of bacteria that degraded ceftiofur belonged to the genera Bacillus and Bacteroides. PCR analysis of bacterial DNA detected specific β-lactamase genes. Bacillus cereus and B. mycoides isolates produced extended-spectrum β-lactamases and metallo-β-lactamases. Seven isolates of Bacteroides spp. produced multiple β-lactamases, including possibly CepA, and metallo-β-lactamases. Isolates of Eubacterium biforme, Bifidobacterium breve, and several Clostridium spp. also produced ceftiofur-degrading β-lactamases. An agar gel overlay technique on isoelectric focusing separations of bacterial lysates showed that β-lactamase enzymes were sufficient to degrade ceftiofur. These results suggest that ceftiofur is inactivated nonenzymatically and degraded enzymatically by multiple β-lactamases from bacteria in the large intestines of cattle.

ALA-based fluorescent diagnosis of malignant oral lesions in the presence of bacterial porphyrin formation

NASA Astrophysics Data System (ADS)

Schleier, P.; Berndt, A.; Zinner, K.; Zenk, W.; Dietel, W.; Pfister, W.

2006-02-01

The aminolevulinic acid (5-ALA) -based fluorescence diagnosis has been found to be promising for an early detection and demarcation of superficial oral squamous cell carcinomas (OSCC). This method has previously demonstrated high sensitivity, however this clinical trial showed a specificity of approximately 62 %. This specificity was mainly restricted by tumor detection in the oral cavity in the presence of bacteria. After topical ALA application in the mouth of patients with previously diagnosed OSSC, red fluorescent areas were observed which did not correlate to confirm histological findings. Swabs and plaque samples were taken from 44 patients and cultivated microbiologically. Fluorescence was investigated (OMA-system) from 32 different bacteria strains found naturally in the oral cavity. After ALA incubation, 30 of 32 strains were found to synthesize fluorescent porphyrins, mainly Protoporphyrin IX. Also multiple fluorescent spectra were obtained having peak wavelengths of 636 nm and around 618 nm - 620 nm indicating synthesis of different porphyrins, such as the lipophylic Protoporphyrin IX (PpIX) and hydrophylic porphyrins (water soluble porphyrins, wsp). Of the 32 fluorescent bacterial strains, 18 produced wsp, often in combination with PpIX, and 5 produced solely wsp. These results clarify that ALA-based fluorescence diagnosis without consideration or suppression of bacteria fluorescence may lead to false-positive findings. It is necessary to suppress bacteria fluorescence with suitable antiseptics before starting the procedure. In this study, when specific antiseptic pre-treatment was performed bacterial associated fluorescence was significantly reduced.

An efficient tag derived from the common epitope of tospoviral NSs proteins for monitoring recombinant proteins expressed in both bacterial and plant systems.

PubMed

Cheng, Hao-Wen; Chen, Kuan-Chun; Raja, Joseph A J; Li, Jian-Xian; Yeh, Shyi-Dong

2013-04-15

NSscon (23 aa), a common epitope in the gene silencing suppressor NSs proteins of the members of the Watermelon silver mottle virus (WSMoV) serogroup, was previously identified. In this investigation, we expressed different green fluorescent protein (GFP)-fused deletions of NSscon in bacteria and reacted with NSscon monoclonal antibody (MAb). Our results indicated that the core 9 amino acids, "(109)KFTMHNQIF(117)", denoted as "nss", retain the reactivity of NSscon. In bacterial pET system, four different recombinant proteins labeled with nss, either at N- or C-extremes, were readily detectable without position effects, with sensitivity superior to that for the polyhistidine-tag. When the nss-tagged Zucchini yellow mosaic virus (ZYMV) helper component-protease (HC-Pro) and WSMoV nucleocapsid protein were transiently expressed by agroinfiltration in tobacco, they were readily detectable and the tag's possible efficacy for gene silencing suppression was not noticed. Co-immunoprecipitation of nss-tagged and non-tagged proteins expressed from bacteria confirmed the interaction of potyviral HC-Pro and coat protein. Thus, we conclude that this novel nss sequence is highly valuable for tagging recombinant proteins in both bacterial and plant expression systems. Copyright © 2013 Elsevier B.V. All rights reserved.

Olfactory Nerve—A Novel Invasion Route of Neisseria meningitidis to Reach the Meninges

PubMed Central

Sjölinder, Hong; Jonsson, Ann-Beth

2010-01-01

Neisseria meningitidis is a human-specific pathogen with capacity to cause septic shock and meningitis. It has been hypothesized that invasion of the central nervous system (CNS) is a complication of a bacteremic condition. In this study, we aimed to characterize the invasion route of N. meningitidis to the CNS. Using an intranasally challenged mouse disease model, we found that twenty percent of the mice developed lethal meningitis even though no bacteria could be detected in blood. Upon bacterial infection, epithelial lesions and redistribution of intracellular junction protein N-cadherin were observed at the nasal epithelial mucosa, especially at the olfactory epithelium, which is functionally and anatomically connected to the CNS. Bacteria were detected in the submucosa of the olfactory epithelium, along olfactory nerves in the cribriform plate, at the olfactory bulb and subsequently at the meninges and subarachnoid space. Furthermore, our data suggest that a threshold level of bacteremia is required for the development of meningococcal sepsis. Taken together, N. meningitidis is able to pass directly from nasopharynx to meninges through the olfactory nerve system. This study enhances our understanding how N. meningitidis invades the meninges. The nasal olfactory nerve system may be a novel target for disease prevention that can improve outcome and survival. PMID:21124975

Olfactory nerve--a novel invasion route of Neisseria meningitidis to reach the meninges.

PubMed

Sjölinder, Hong; Jonsson, Ann-Beth

2010-11-18

Neisseria meningitidis is a human-specific pathogen with capacity to cause septic shock and meningitis. It has been hypothesized that invasion of the central nervous system (CNS) is a complication of a bacteremic condition. In this study, we aimed to characterize the invasion route of N. meningitidis to the CNS. Using an intranasally challenged mouse disease model, we found that twenty percent of the mice developed lethal meningitis even though no bacteria could be detected in blood. Upon bacterial infection, epithelial lesions and redistribution of intracellular junction protein N-cadherin were observed at the nasal epithelial mucosa, especially at the olfactory epithelium, which is functionally and anatomically connected to the CNS. Bacteria were detected in the submucosa of the olfactory epithelium, along olfactory nerves in the cribriform plate, at the olfactory bulb and subsequently at the meninges and subarachnoid space. Furthermore, our data suggest that a threshold level of bacteremia is required for the development of meningococcal sepsis. Taken together, N. meningitidis is able to pass directly from nasopharynx to meninges through the olfactory nerve system. This study enhances our understanding how N. meningitidis invades the meninges. The nasal olfactory nerve system may be a novel target for disease prevention that can improve outcome and survival.

[Development and comparative evaluation of up-converting phosphor technology based lateral flow assay for rapid detection of Yersinia pestis, Bacillus anthracis spore and Brucella spp].

PubMed

Li, Chunfeng; Zhang, Pingping; Wang, Xiaoying; Liu, Xiao; Zhao, Yong; Sun, Chongyun; Wang, Chengbin; Yang, Ruifu; Zhou, Lei

2015-01-01

To develop an up-converting phosphor technology based lateral flow (UPT-LF) assay for rapid and quantitative detection of Yersinia pestis, Bacillus anthracis spore and Brucella spp.and make the comparison with BioThreat Alert (BTA) test strips (Tetracore Inc., USA). Using up-converting phosphor nano-particles (UCP-NPs) as the bio-marker, three double-antibody-sandwich model based UPT-LF strips including Plague-UPT-LF, Anthrax-UPT-LF, Brucella-UPT-LF were prepared and its sensitivity, accuracy, linearity and specificity were determined by detecting 10(10), 10(9), 10(8), 10(7), 10(6), 10(5) and 0 CFU/ml series of concentrations of Y.pestis, B.anthracis, Brucella standards and other 27 kinds of 10(9) CFU/ml series of contrations of bacteria strains.Furthermore, the speed, sensitivity and accuracy of bacteria standards and simulated sample detection were compared between UPT-LF and BTA system. The detection limit of Plague-UPT-LF, Anthrax-UPT-LF and Brucella-LF was 10(5) CFU/ml. The CV of series of bacteria concentrations was ≤ 15%, and the r between lg (T/C-cut-off) and lg (concentration) was 0.996,0.998 and 0.999 (F values were 1 647.57, 743.51 and 1 822.17. All the P values were <0.001), respectively. The specificity of Plague-UPT-LF and Brucella-LF were excellent, while that of Anthrax-UPT-LF was a little bit regretful because of non-specific reaction with two isolates of B. subtilis and one B.cereus. On-site evaluation showed the detection time of UPT-LF for all Y.pestis, B.anthracis spore and Brucella spp.was 33, 36 and 37 min, while BTA was 115, 115 and 111 min, which revealed the higher detection speed and sensitivity of UPT-LF comparing with BTA. The negative rate of two methods for blank standard was both 5/5, the sensitivity of UPT-LF for Y.pestis,B.anthracis spore and Brucella spp. was all 10(5) CFU/ml, then BTA was 10(6), 10(6) and 10(5) CFU/ml, respectively. The detection rate of UPT-LF for all three bacteria analog positive samples was 16/16, while BTA for B.anthracis was 7/16 only. The good performance including rapidness, simplicity and high sensitivity will bring the bright future of UPT-LF to be broadly used on-site as first response to bio-terrorism.

The occurrence of pathogenic bacteria in some ships' ballast water incoming from various marine regions to the Sea of Marmara, Turkey.

PubMed

Altug, Gulsen; Gurun, Sevan; Cardak, Mine; Ciftci, Pelin S; Kalkan, Samet

2012-10-01

The composition and frequency of antibiotic resistance of pathogenic bacteria, the abundance of heterotrophic aerobic bacteria (HPC) and possible in-situ use of chromogenic agar were investigated in the ships' ballast water coming from different regions of the world to the Sea of Marmara, Turkey for the first time. The samples that were taken from 21 unit ships coming from various marine environments of the Southern China Sea, the Atlantic Ocean, the Mediterranean and the Black Sea to the Sea of Marmara, Turkey in 2009 and 2010 were tested. 38 bacteria species, 27 of them pathogenic bacteria belonging to 17 familia, were detected. Vibrio cholera was not detected in the samples. However, the presence of a high number of HPC, including a cocktail of pathogenic bacteria showed that the ships carry a potential risk for the Sea of Marmara. Copyright © 2012 Elsevier Ltd. All rights reserved.

Symptomatic and asymptomatic apical periodontitis associated with red complex bacteria: clinical and microbiological evaluation.

PubMed

Buonavoglia, Alessio; Latronico, Francesca; Pirani, Chiara; Greco, Maria Fiorella; Corrente, Marialaura; Prati, Carlo

2013-01-01

In this study, the association of red complex (RC) bacteria that include Treponema denticola, Tannerella forsythia and Porphyromonas gingivalis with acute, exacerbated or chronic apical periodontitis was evaluated. Seventy-one patients with periapical disease were evaluated by clinical examination and microbiological samples obtained from the root canals were analyzed by a polymerase chain reaction assay. Twenty-one (29.6%) samples were positive for RC bacteria, with T. denticola, T. forsythia and P. gingivalis being detected in 14 (19.7%), 10 (14.1%) and 6 (8.5%) samples, respectively. RC bacteria were mainly associated with acute apical periodontitis (29.2%) and phoenix abscess (63.2%), while they were only sporadically detected (7.1%) in patients with chronic apical periodontitis. Generally, RC bacteria were associated with pain and a higher frequency of intracanalar/intrasulcular pus drainage. Involvement of RC bacteria in symptomatic periapical disease should be suspected in the presence of particularly severe clinical pain and pus drainage.

The extended spectrum β-lactamases (ESBL) and virulence genes of intestinal enteroaggregative Escherichia coli (EAEC) in healthy elderly individuals.

PubMed

Wang, Yuan; Wu, Jian; Cao, Yi

2015-01-01

to analyze the detection rate of intestinal enteroaggregative Escherichia coli (EAEC) in healthy elderly (≥60 years) individuals in the Hangzhou area of China, and to investigate the extended spectrum β-lactamases and virulence genes of EAEC. Stool specimens provided by healthy elderly individuals were cultured on blood agar, SS, and MAC plates. The bacterial strains were identified using Vitek-2 Compact automatic microorganism identification system and mass spectrometry. The resistance phenotypes of the bacteria were determined using the double-disk synergy method. The resistance genes and the EAEC virulence gene, astA and aggR, were amplified by PCR and compared to the sequences available in Gen Bank. Among the 1050 healthy volunteers, the majority of bacteria were E. coli, accounting for 960 strains, with an ESBL-positive rate of 36.3% (348/960). The EAEC detection rate was 10% (96/960); among them, 84 strains were astA, the detection rate of which was 8.75%; 12 strains were aggR, the detection rate of which was 1.25%. The ESBL-positive rate of EAEC strains were 56.25% (54/96), all of which carried the CTX-M type, with the CTX-M-14 predominating at 66.7% (36/54). The ESBL-positive rate of intestinal E. coli in healthy elderly individuals in the Hangzhou area of China was higher than the rate detected in other regions of china; and there was a high rate of antibiotic resistance among the intestinal EAEC in healthy elderly individuals. The results of this study suggest that EAEC is not only a pathogenic bacteria detected in diarrhea patients, but can also be present in healthy individuals, and high-resistance clinical strains have spread to the healthy population in the Hangzhou area. So vigilance is critical.

Six years' experience of using the BacT/ALERT system to screen all platelet concentrates, and additional testing of outdated platelet concentrates to estimate the frequency of false-negative results.

PubMed

Larsen, C P; Ezligini, F; Hermansen, N O; Kjeldsen-Kragh, J

2005-02-01

Approximately 1 in every 2000 units of platelets is contaminated with bacteria. The BacT/ALERT automated blood culture system can be used to screen platelet concentrates (PCs) for bacterial contamination. Data were collected from May 1998 until May 2004. The number of PCs tested during this period was 36 896, most of which were produced from pools of four buffy-coats. On the day following blood collection or platelet apheresis, a 5-10 ml sample of the PC was aseptically transferred to a BacT/ALERT culture bottle for detection of aerobic bacteria. The sample was monitored for bacterial growth during the entire storage period of the PC (6.5 days). When a positive signal was generated, the culture bottle, the PC and the erythrocyte concentrates were tested for bacterial growth. In order to determine the frequency of false-negative BacT/ALERT signals, 1061 outdated PCs were tested during the period from May 2002 to May 2004. Eighty-eight positive signals were detected by the BacT/ALERT system, of which 12 were interpreted as truly positive. Fourteen signals were interpreted as truly false positive. Thirty-three signals were interpreted to be probably false positive. Two of 1061 outdated units tested positive, and Bacillus spp. and Staphylococcus epidermidis, respectively, were isolated from these PCs. Between 0.03% and 0.12% of the PCs were contaminated with bacteria. BacT/ALERT is an efficient tool for monitoring PCs for bacterial contamination; however, it is important to realize that false-negative results may occur.

Frequency domain fluorescence lifetime imaging microscopy system for detecting inflammatory cells

NASA Astrophysics Data System (ADS)

Yahav, Gilad; Gershanov, Sivan; Salmon-Divon, Mali; Ben-Zvi, Haim; Mircus, Gabriel; Goldenberg-Cohen, Nitza; Fixler, Dror

2018-02-01

Characterizing different pathological states in the cellular level with a high throughput diagnostic tool is one of the main interests today. In previously works, we demonstrated how the frequency domain (FD) fluorescence lifetime imaging microscopy (FLIM) technique could be utilized to implement that in variety of examples. Among them was to classify between different chromosomal abnormalities in patients with b-cell chronic lymphocytic leukemia (B-CLL) and between metastatic cells and inflammation cells in the cerebral spinal fluid of patients with Medulloblastoma. This research describes the use of FD-FLIM system to differentiate between patients diagnosed without any disease (controls) that showed a normal median FLT (2.65+/-0.11ns) and patients diagnosed with inflammation (viruses and bacteria) that showed a prolong median FLT and a larger distribution (3.18+/-0.44ns in viruses and 3.28+/-0.45ns). The study group of this research included 43 samples divided into 4 groups: 9 samples diagnosed with different types of bacteria, 16 samples diagnosed with different types of viruses, 12 samples diagnosed with no any bacteria or virus and 5 samples diagnosed without any disease that served as controls. Furthermore, we studied a group of patients without detection of inflammation that were sick. We found that this group was divided into two groups; one group had the same median FLT as the controls, and the other group had the same median FLT as the inflammatory patients. As a result, we believe the FD-FLIM system can suggest a faster and more accurate diagnostic technique than the methods used today. The correlations of the FLT distribution pattern with the different groups are presented.

Bacteriological quality of vegetables from organic and conventional production in different areas of Korea.

PubMed

Tango, Charles Nkufi; Choi, Na-Jung; Chung, Myung-Sub; Oh, Deog Hwan

2014-08-01

Foods grown in organic production systems have been described as representing an increased risk to public health compared with foods from conventional production. Leafy vegetables (spinach, romaine lettuce, and green sesame leaves) grown in organic and conventional systems were collected from various areas in Korea and examined using standard culture methods to compare the microbiological quality of the produce grown in the two agricultural systems. The 354 samples of these leafy vegetables were analyzed for levels of indicator bacteria (aerobic bacteria, coliforms, and Escherichia coli) and the prevalence of the pathogens Staphylococcus aureus, E. coli O157:H7, Listeria monocytogenes, Bacillus cereus, and Salmonella. Aerobic bacteria and coliforms were detected in all vegetable types, but nonpathogenic E. coli was below the limit of detection in all samples. B. cereus was the most prevalent pathogen, found on 7 (11.1%) of the 63 organic spinach samples. The prevalence of S. aureus was highest in organic sesame leaves; it was found on 5 (8.0%) of the 63 samples. The prevalence of L. monocytogenes was highest on organic romaine lettuce and spinach; it was found in 4 (6.4%) of 63 samples of each type of vegetable. E. coli O157:H7 found on only 1 (1.58%) of 55 conventional spinach samples. These results suggest that farming type at most only slightly affects the hygienic quality of leafy vegetables, and no effect was found for sample collection area. Salmonella was not isolated from any of the conventional or organic leafy vegetables. These results do not support the hypothesis that organic produce poses a substantially greater risk of pathogen contamination than does conventional produce.

[Comparison between rapid detection method of enzyme substrate technique and multiple-tube fermentation technique in water coliform bacteria detection].

PubMed

Sun, Zong-ke; Wu, Rong; Ding, Pei; Xue, Jin-Rong

2006-07-01

To compare between rapid detection method of enzyme substrate technique and multiple-tube fermentation technique in water coliform bacteria detection. Using inoculated and real water samples to compare the equivalence and false positive rate between two methods. Results demonstrate that enzyme substrate technique shows equivalence with multiple-tube fermentation technique (P = 0.059), false positive rate between the two methods has no statistical difference. It is suggested that enzyme substrate technique can be used as a standard method for water microbiological safety evaluation.

Ammonia- and nitrite-oxidizing bacterial communities in a pilot-scale chloraminated drinking water distribution system.

PubMed

Regan, John M; Harrington, Gregory W; Noguera, Daniel R

2002-01-01

Nitrification in drinking water distribution systems is a common operational problem for many utilities that use chloramines for secondary disinfection. The diversity of ammonia-oxidizing bacteria (AOB) and nitrite-oxidizing bacteria (NOB) in the distribution systems of a pilot-scale chloraminated drinking water treatment system was characterized using terminal restriction fragment length polymorphism (T-RFLP) analysis and 16S rRNA gene (ribosomal DNA [rDNA]) cloning and sequencing. For ammonia oxidizers, 16S rDNA-targeted T-RFLP indicated the presence of Nitrosomonas in each of the distribution systems, with a considerably smaller peak attributable to Nitrosospira-like AOB. Sequences of AOB amplification products aligned within the Nitrosomonas oligotropha cluster and were closely related to N. oligotropha and Nitrosomonas ureae. The nitrite-oxidizing communities were comprised primarily of Nitrospira, although Nitrobacter was detected in some samples. These results suggest a possible selection of AOB related to N. oligotropha and N. ureae in chloraminated systems and demonstrate the presence of NOB, indicating a biological mechanism for nitrite loss that contributes to a reduction in nitrite-associated chloramine decay.

Ammonia- and Nitrite-Oxidizing Bacterial Communities in a Pilot-Scale Chloraminated Drinking Water Distribution System

PubMed Central

Regan, John M.; Harrington, Gregory W.; Noguera, Daniel R.

2002-01-01

Nitrification in drinking water distribution systems is a common operational problem for many utilities that use chloramines for secondary disinfection. The diversity of ammonia-oxidizing bacteria (AOB) and nitrite-oxidizing bacteria (NOB) in the distribution systems of a pilot-scale chloraminated drinking water treatment system was characterized using terminal restriction fragment length polymorphism (T-RFLP) analysis and 16S rRNA gene (ribosomal DNA [rDNA]) cloning and sequencing. For ammonia oxidizers, 16S rDNA-targeted T-RFLP indicated the presence of Nitrosomonas in each of the distribution systems, with a considerably smaller peak attributable to Nitrosospira-like AOB. Sequences of AOB amplification products aligned within the Nitrosomonas oligotropha cluster and were closely related to N. oligotropha and Nitrosomonas ureae. The nitrite-oxidizing communities were comprised primarily of Nitrospira, although Nitrobacter was detected in some samples. These results suggest a possible selection of AOB related to N. oligotropha and N. ureae in chloraminated systems and demonstrate the presence of NOB, indicating a biological mechanism for nitrite loss that contributes to a reduction in nitrite-associated chloramine decay. PMID:11772611

Cultural and Molecular Evidence of Legionella spp. Colonization in Dental Unit Waterlines: Which Is the Best Method for Risk Assessment?

PubMed

Ditommaso, Savina; Giacomuzzi, Monica; Ricciardi, Elisa; Zotti, Carla M

2016-02-06

Legionella spp. are ubiquitous in aquatic habitats and water distribution systems, including dental unit waterlines (DUWLs). The aim of the present study was to determine the prevalence of Legionella in DUWLs and tap water samples using PMA-qPCR and standard culture methods. The total viable counts (TVCs) of aerobic heterotrophic bacteria in the samples were also determined. Legionella spp. were detected and quantified using the modified ISO 11731 culture method. Extracted genomic DNA was analysed using the iQ-Check Quanti Legionella spp. kit, and the TVCs were determined according to the ISO protocol 6222. Legionella spp. were detected in 100% of the samples using the PMA-qPCR method, whereas these bacteria were detected in only 7% of the samples using the culture method. The number of colony forming units (CFUs) of the TVCs in the DUWL and tap water samples differed, with the bacterial load being significantly lower in the tap water samples (p-value = 0). The counts obtained were within the Italian standard range established for potable water in only 5% of the DUWL water samples and in 77% of the tap water samples. Our results show that the level of Legionella spp. contamination determined using the culture method does not reflect the true scale of the problem, and consequently we recommend testing for the presence of aerobic heterotrophic bacteria based on the assumption that Legionella spp. are components of biofilms.

Cultural and Molecular Evidence of Legionella spp. Colonization in Dental Unit Waterlines: Which Is the Best Method for Risk Assessment?

PubMed Central

Ditommaso, Savina; Giacomuzzi, Monica; Ricciardi, Elisa; Zotti, Carla M.

2016-01-01

Legionella spp. are ubiquitous in aquatic habitats and water distribution systems, including dental unit waterlines (DUWLs). The aim of the present study was to determine the prevalence of Legionella in DUWLs and tap water samples using PMA-qPCR and standard culture methods. The total viable counts (TVCs) of aerobic heterotrophic bacteria in the samples were also determined. Legionella spp. were detected and quantified using the modified ISO 11731 culture method. Extracted genomic DNA was analysed using the iQ-Check Quanti Legionella spp. kit, and the TVCs were determined according to the ISO protocol 6222. Legionella spp. were detected in 100% of the samples using the PMA-qPCR method, whereas these bacteria were detected in only 7% of the samples using the culture method. The number of colony forming units (CFUs) of the TVCs in the DUWL and tap water samples differed, with the bacterial load being significantly lower in the tap water samples (p-value = 0). The counts obtained were within the Italian standard range established for potable water in only 5% of the DUWL water samples and in 77% of the tap water samples. Our results show that the level of Legionella spp. contamination determined using the culture method does not reflect the true scale of the problem, and consequently we recommend testing for the presence of aerobic heterotrophic bacteria based on the assumption that Legionella spp. are components of biofilms. PMID:26861373

Instrument Would Detect and Collect Biological Aerosols

NASA Technical Reports Server (NTRS)

Savoy, Steve; Mayo, Mike

2006-01-01

A proposed compact, portable instrument would sample micron-sized airborne particles, would discriminate between biological ones (e.g., bacteria) and nonbiological ones (e.g., dust particles), and would collect the detected biological particles for further analysis. The instrument is intended to satisfy a growing need for means of rapid, inexpensive collection of bioaerosols in a variety of indoor and outdoor settings. Purposes that could be served by such collection include detecting airborne pathogens inside buildings and their ventilation systems, measuring concentrations of airborne biological contaminants around municipal waste-processing facilities, monitoring airborne effluents from suspected biowarfare facilities, and warning of the presence of airborne biowarfare agents

New biosensors for food safety screening solutions

NASA Astrophysics Data System (ADS)

Dyer, Maureen A.; Oberholtzer, Jennifer A.; Mulligan, David C.; Hanson, William P.

2009-05-01

Hanson Technologies has developed the automated OmniFresh 1000 system to sample large volumes of produce wash water, collect the pathogens, and detect their presence. By collecting a continuous sidestream of wash water, the OmniFresh uses a sample that represent the entire lot of produce being washed. The OmniFresh does not require bacterial culture or enrichment, and it detects both live and dead bacteria in the collected sample using an in-line sensor. Detection occurs in an array biosensor capable of handling large samples with complex matrices. Additionally, sample can be sent for traditional confirming tests after the screening performed by the OmniFresh.

Silkworm larvae plasma (SLP) assay for detection of bacteria: False positives secondary to inflammation in vivo.

PubMed

Ma, Michelle; Rice, Tyler A; Percopo, Caroline M; Rosenberg, Helene F

2017-01-01

The silkworm larvae plasma (SLP) assay has been developed as a means to detect bacterial peptidoglycan as a surrogate for live bacteria. Here, we present results that indicate that generation of melanin by this assay is not fully reliable as a surrogate marker for bacterial count. Published by Elsevier B.V.

9 CFR 113.26 - Detection of viable bacteria and fungi except in live vaccine.

Code of Federal Regulations, 2012 CFR

2012-01-01

... 9 Animals and Animal Products 1 2012-01-01 2012-01-01 false Detection of viable bacteria and fungi except in live vaccine. 113.26 Section 113.26 Animals and Animal Products ANIMAL AND PLANT HEALTH... in live vaccine. Each serial and subserial of biological product except live vaccines shall be tested...

9 CFR 113.26 - Detection of viable bacteria and fungi except in live vaccine.

Code of Federal Regulations, 2014 CFR

2014-01-01

... 9 Animals and Animal Products 1 2014-01-01 2014-01-01 false Detection of viable bacteria and fungi except in live vaccine. 113.26 Section 113.26 Animals and Animal Products ANIMAL AND PLANT HEALTH... in live vaccine. Each serial and subserial of biological product except live vaccines shall be tested...

9 CFR 113.26 - Detection of viable bacteria and fungi except in live vaccine.

Code of Federal Regulations, 2013 CFR

2013-01-01

... 9 Animals and Animal Products 1 2013-01-01 2013-01-01 false Detection of viable bacteria and fungi except in live vaccine. 113.26 Section 113.26 Animals and Animal Products ANIMAL AND PLANT HEALTH... in live vaccine. Each serial and subserial of biological product except live vaccines shall be tested...

Air sampling to assess potential generation of aerosolized viable bacteria during flow cytometric analysis of unfixed bacterial suspensions

PubMed Central

Carson, Christine F; Inglis, Timothy JJ

2018-01-01

This study investigated aerosolized viable bacteria in a university research laboratory during operation of an acoustic-assisted flow cytometer for antimicrobial susceptibility testing by sampling room air before, during and after flow cytometer use. The aim was to assess the risk associated with use of an acoustic-assisted flow cytometer analyzing unfixed bacterial suspensions. Air sampling in a nearby clinical laboratory was conducted during the same period to provide context for the existing background of microorganisms that would be detected in the air. The three species of bacteria undergoing analysis by flow cytometer in the research laboratory were Klebsiella pneumoniae, Burkholderia thailandensis and Streptococcus pneumoniae. None of these was detected from multiple 1000 L air samples acquired in the research laboratory environment. The main cultured bacteria in both locations were skin commensal and environmental bacteria, presumed to have been disturbed or dispersed in laboratory air by personnel movements during routine laboratory activities. The concentrations of bacteria detected in research laboratory air samples were reduced after interventional cleaning measures were introduced and were lower than those in the diagnostic clinical microbiology laboratory. We conclude that our flow cytometric analyses of unfixed suspensions of K. pneumoniae, B. thailandensis and S. pneumoniae do not pose a risk to cytometer operators or other personnel in the laboratory but caution against extrapolation of our results to other bacteria and/or different flow cytometric experimental procedures. PMID:29608197

Using Raman spectroscopy and SERS for in situ studies of rhizosphere bacteria

DOE Office of Scientific and Technical Information (OSTI.GOV)

Mohseni, Hooman; Agahi, Massoud H.; Razeghi, Manijeh

Bacteria colonize plant roots to form a symbiotic relationship with the plant and can play in important role in promoting plant growth. Raman spectroscopy is a useful technique to study these bacterial systems and the chemical signals they utilize to interact with the plant. We present a Raman study of Pantoea YR343 that was isolated from the rhizosphere of Populus deltoides (Eastern Cottonwood). Pantoea sp. YR343 produce yellowish carotenoid pigment that play a role in protection against UV radiation, in the anti-oxidative pathways and in membrane fluidity. Raman spectroscopy is used to non-invasively characterize the membrane bound carotenoids. The spectramore » collected from a mutant strain created by knocking out the crtB gene that encodes a phytoene synthase responsible for early stage of carotenoid biosynthesis, lack the carotenoid peaks. Surface Enhanced Raman Spectroscopy is being employed to detect the plant phytoharmone indoleacetic acid that is synthesized by the bacteria. This work describes our recent progress towards utilizing Raman spectroscopy as a label free, non-destructive method of studying plant-bacteria interactions in the rhizosphere.« less

Co-relation of estrous cycle phases with uterine bacterial and fungal flora in non-pregnant female laboratory rabbits

PubMed Central

Mogheiseh, A.; Derakhshandeh, A.; Batebi, E.; Golestani, N.; Moshiri, A.

2017-01-01

This study was designed to investigate the relationship between the estrous cycle phases with uterine bacterial and fungal flora in non-pregnant female rabbits. Thirty laboratory mature multiparous rabbits were used for this purpose. Samples from uterine lavage for culture of bacteria and fungi were collected at different stages of estrous cycle (based on vaginal cytology), and histopathological observations were evaluated based on the scoring system used for defining the infection of the uterus. Various types of bacteria and fungi were isolated from rabbits at all stages of estrous cycle. The widest variety of bacteria and fungi was isolated at Di-estrous stage and the lowest variety was detected at estrous stage. Klebsiella oxytoca as well as yeast have been isolated at all stages of estrous cycle. This study showed that infection with K. oxytoca and yeast had no relationship with different stages of estrous cycle but other bacteria and fungus were associated with one or more stages of the estrous cycle in rabbits. PMID:28775754

Assessment of drinking water quality using indicator bacteria and bacteriophages.

PubMed

Méndez, Javier; Audicana, Ana; Cancer, Mercedes; Isern, Anna; Llaneza, Julian; Moreno, Belén; Navarro, Mercedes; Tarancón, M Lluisa; Valero, Fernando; Ribas, Ferran; Jofre, Juan; Lucena, Francisco

2004-09-01

Bacterial indicators and bacteriophages suggested as potential indicators of water quality were determined by public laboratories in water from springs, household water wells, and rural and metropolitan water supplies in north-eastern Spain. Indicator bacteria were detected more frequently than bacteriophages in springs, household water wells and rural water supplies. In contrast, positive bacteriophage detections were more numerous than those of bacteria in metropolitan water supplies. Most of the metropolitan water supply samples containing indicators had concentrations of chlorine below 0.1 mg l(-1), their indicator loads resembling more closely those of rural water supplies than any other samples taken from metropolitan water supplies. The number of samples from metropolitan water supplies containing more than 0.1 mg l(-1) of chlorine that contained phages clearly outnumbered those containing indicator bacteria. Some association was observed between rainfall and the presence of indicators. Sediments from service reservoirs and water from dead ends in the distribution network of one of the metropolitan water supplies were also tested. Bacterial indicators and phages were detected in a higher percentage than in samples of tap water from the same network. Additionally, indicator bacteria were detected more frequently than bacteriophages in sediments of service reservoirs and water from dead end samples. We conclude that naturally occurring indicator bacteria and bacteriophages respond differently to chlorination and behave differently in drinking water distribution networks. Moreover, this study has shown that testing for the three groups of phages in routine laboratories is easy to implement and feasible without the requirement for additional material resources for the laboratories.

A Simple Method for Assessment of MDR Bacteria for Over-Expressed Efflux Pumps

PubMed Central

Martins, Marta; McCusker, Matthew P; Viveiros, Miguel; Couto, Isabel; Fanning, Séamus; Pagès, Jean-Marie; Amaral, Leonard

2013-01-01

It is known that bacteria showing a multi-drug resistance phenotype use several mechanisms to overcome the action of antibiotics. As a result, this phenotype can be a result of several mechanisms or a combination of thereof. The main mechanisms of antibiotic resistance are: mutations in target genes (such as DNA gyrase and topoisomerase IV); over-expression of efflux pumps; changes in the cell envelope; down regulation of membrane porins, and modified lipopolysaccharide component of the outer cell membrane (in the case of Gram-negative bacteria). In addition, adaptation to the environment, such as quorum sensing and biofilm formation can also contribute to bacterial persistence. Due to the rapid emergence and spread of bacterial isolates showing resistance to several classes of antibiotics, methods that can rapidly and efficiently identify isolates whose resistance is due to active efflux have been developed. However, there is still a need for faster and more accurate methodologies. Conventional methods that evaluate bacterial efflux pump activity in liquid systems are available. However, these methods usually use common efflux pump substrates, such as ethidium bromide or radioactive antibiotics and therefore, require specialized instrumentation, which is not available in all laboratories. In this review, we will report the results obtained with the Ethidium Bromide-agar Cartwheel method. This is an easy, instrument-free, agar based method that has been modified to afford the simultaneous evaluation of as many as twelve bacterial strains. Due to its simplicity it can be applied to large collections of bacteria to rapidly screen for multi-drug resistant isolates that show an over-expression of their efflux systems. The principle of the method is simple and relies on the ability of the bacteria to expel a fluorescent molecule that is substrate for most efflux pumps, ethidium bromide. In this approach, the higher the concentration of ethidium bromide required to produce fluorescence of the bacterial mass, the greater the efflux capacity of the bacterial cells. We have tested and applied this method to a large number of Gram-positive and Gram-negative bacteria to detect efflux activity among these multi-drug resistant isolates. The presumptive efflux activity detected by the Ethidium Bromide-agar Cartwheel method was subsequently confirmed by the determination of the minimum inhibitory concentration for several antibiotics in the presence and absence of known efflux pump inhibitors. PMID:23589748

A Rapid and Simple Real-Time PCR Assay for Detecting Foodborne Pathogenic Bacteria in Human Feces.

PubMed

Hanabara, Yutaro; Ueda, Yutaka

2016-11-22

A rapid, simple method for detecting foodborne pathogenic bacteria in human feces is greatly needed. Here, we examined the efficacy of a method that employs a combination of a commercial PCR master mix, which is insensitive to PCR inhibitors, and a DNA extraction method which used sodium dodecyl benzene sulfonate (SDBS), and Tween 20 to counteract the inhibitory effects of SDBS on the PCR assay. This method could detect the target genes (stx1 and stx2 of enterohemorrhagic Escherichia coli, invA of Salmonella Enteritidis, tdh of Vibrio parahaemolyticus, gyrA of Campylobacter jejuni, ceuE of Campylobacter coli, SEA of Staphylococcus aureus, ces of Bacillus cereus, and cpe of Clostridium perfringens) in a fecal suspension containing 1.0 × 10 1 to 1.0 × 10 3 CFU/ml. Furthermore, the assay was neither inhibited nor influenced by individual differences among the fecal samples of 10 subjects or fecal concentration (40-160 mg/ml in the fecal suspension). When we attempted to detect the genes of pathogenic bacteria in 4 actual clinical cases, we found that this method was more sensitive than standard culture method. These results showed that this assay is a rapid, simple detection method for foodborne pathogenic bacteria in human feces.

Use of Whole-Cell Bioassays for Screening Quorum Signaling, Quorum Interference, and Biofilm Dispersion.

PubMed

Thornhill, Starla G; McLean, Robert J C

2018-01-01

In most bacteria, a global level of regulation, termed quorum sensing (QS), exists involving intercellular communication via the production and response to cell density-dependent signal molecules. QS has been associated with a number of important features in bacteria including virulence regulation and biofilm formation. Consequently, there is considerable interest in understanding, detecting, and inhibiting QS. N-acylated homoserine lactones (AHLs) are used as extracellular QS signals by a variety of Gram-negative bacteria. Chromobacterium violaceum, commonly found in soil and water, produces the characteristic purple pigment violacein, regulated by AHL-mediated QS. Based on this readily observed pigmentation phenotype, C. violaceum strains can be used to detect various aspects of AHL-mediated QS activity. In another commonly used bioassay organism, Agrobacterium tumefaciens, QS can be detected by the use of a reporter gene such as lacZ. Here, we describe several commonly used approaches incorporating C. violaceum and A. tumefaciens that can be used to detect AHL and QS inhibitors. Due to the inherent low susceptibility of biofilm bacteria to antimicrobial agents, biofilm dispersion, whereby bacteria reenter the planktonic community, is another increasingly important area of research. At least one signal, distinct from traditional QS, has been identified and there are a variety of other environmental factors that also trigger dispersion. We describe a microtiter-based experimental strategy whereby potential biofilm dispersion compounds can be screened.

Rapid Spoligotyping of Mycobacterium tuberculosis Complex Bacteria by Use of a Microarray System with Automatic Data Processing and Assignment

PubMed Central

Ruettger, Anke; Nieter, Johanna; Skrypnyk, Artem; Engelmann, Ines; Ziegler, Albrecht; Moser, Irmgard; Monecke, Stefan; Ehricht, Ralf

2012-01-01

Membrane-based spoligotyping has been converted to DNA microarray format to qualify it for high-throughput testing. We have shown the assay's validity and suitability for direct typing from tissue and detecting new spoligotypes. Advantages of the microarray methodology include rapidity, ease of operation, automatic data processing, and affordability. PMID:22553239

Rapid spoligotyping of Mycobacterium tuberculosis complex bacteria by use of a microarray system with automatic data processing and assignment.

PubMed

Ruettger, Anke; Nieter, Johanna; Skrypnyk, Artem; Engelmann, Ines; Ziegler, Albrecht; Moser, Irmgard; Monecke, Stefan; Ehricht, Ralf; Sachse, Konrad

2012-07-01

Membrane-based spoligotyping has been converted to DNA microarray format to qualify it for high-throughput testing. We have shown the assay's validity and suitability for direct typing from tissue and detecting new spoligotypes. Advantages of the microarray methodology include rapidity, ease of operation, automatic data processing, and affordability.

Time-Resolved Microfluorescence In Biomedical Diagnosis

NASA Astrophysics Data System (ADS)

Schneckenburger, Herbert

1985-02-01

A measuring system combining subnanosecond laser-induced fluorescence with microscopic signal detection was installed and used for diverse projects in the biomedical and environmental field. These projects are ranging from tumor diagnosis and enzymatic analysis to measurements of the activity of methanogenic bacteria which effect biogas production and waste water cleaning. The advantages of this method and its practical applicability are discussed.

MALDI-TOF mass spectrometry for rapid diagnosis of postoperative endophthalmitis.

PubMed

Mailhac, Adriane; Durand, Harmonie; Boisset, Sandrine; Maubon, Danièle; Berger, Francois; Maurin, Max; Chiquet, Christophe; Bidart, Marie

2017-01-30

This study describes an innovative strategy for rapid detection and identification of bacteria causing endophthalmitis, combining the use of an automated blood culture system with MALDI-TOF mass spectrometry methodology. Using this protocol, we could identify 96% of 45 bacterial strains isolated from vitreous samples collected in acute post-operative endophthalmitis patients. Copyright © 2016 Elsevier B.V. All rights reserved.

Autoradiography and Immunofluorescence Combined for Autecological Study of Single Cell Activity with Nitrobacter as a Model System1

PubMed Central

Fliermans, C. B.; Schmidt, E. L.

1975-01-01

Specific detection of a particular bacterium by immunofluorescence was combined with estimation of its metabolic activity by autoradiography. The nitrifying bacteria Nitrobacter agilis and N. winogradskyi were used as a model system. Nitrobacter were incubated with NaH14CO3 and 14CO2 prior to study. The same preparations made for autoradiograms were stained with fluorescent antibodies specific for the Nitrobacter species. Examination by epifluorescence and transmitted dark-field microscopy revealed Nitrobacter cells with and without associated silver grains. Direct detection and simultaneous evaluation of metabolic activity of Nitrobacter was demonstrated in pure cultures, in a simple mixed culture, and in a natural soil. Images PMID:1103733

Accelerating bacterial growth detection and antimicrobial susceptibility assessment in integrated picoliter droplet platform.

PubMed

Kaushik, Aniruddha M; Hsieh, Kuangwen; Chen, Liben; Shin, Dong Jin; Liao, Joseph C; Wang, Tza-Huei

2017-11-15

There remains an urgent need for rapid diagnostic methods that can evaluate antibiotic resistance for pathogenic bacteria in order to deliver targeted antibiotic treatments. Toward this end, we present a rapid and integrated single-cell biosensing platform, termed dropFAST, for bacterial growth detection and antimicrobial susceptibility assessment. DropFAST utilizes a rapid resazurin-based fluorescent growth assay coupled with stochastic confinement of bacteria in 20 pL droplets to detect signal from growing bacteria after 1h incubation, equivalent to 2-3 bacterial replications. Full integration of droplet generation, incubation, and detection into a single, uninterrupted stream also renders this platform uniquely suitable for in-line bacterial phenotypic growth assessment. To illustrate the concept of rapid digital antimicrobial susceptibility assessment, we employ the dropFAST platform to evaluate the antibacterial effect of gentamicin on E. coli growth. Copyright © 2017 Elsevier B.V. All rights reserved.

Design of a surface-scanning coil detector for direct bacteria detection on food surfaces using a magnetoelastic biosensor

NASA Astrophysics Data System (ADS)

Chai, Yating; Wikle, Howard C.; Wang, Zhenyu; Horikawa, Shin; Best, Steve; Cheng, Zhongyang; Dyer, Dave F.; Chin, Bryan A.

2013-09-01

The real-time, in-situ bacteria detection on food surfaces was achieved by using a magnetoelastic biosensor combined with a surface-scanning coil detector. This paper focuses on the coil design for signal optimization. The coil was used to excite the sensor's vibration and detect its resonant frequency signal. The vibrating sensor creates a magnetic flux change around the coil, which then produces a mutual inductance. In order to enhance the signal amplitude, a theory of the sensor's mutual inductance with the measurement coil is proposed. Both theoretical calculations and experimental data showed that the working length of the coil has a significant effect on the signal amplitude. For a 1 mm-long sensor, a coil with a working length of 1.3 mm showed the best signal amplitude. The real-time detection of Salmonella bacteria on a fresh food surface was demonstrated using this new technology.

Simulation of Graphene Field-Effect Transistor Biosensors for Bacterial Detection.

PubMed

Wu, Guangfu; Meyyappan, Meyya; Lai, King Wai Chiu

2018-05-25

Foodborne illness is correlated with the existence of infectious pathogens such as bacteria in food and drinking water. Probe-modified graphene field effect transistors (G-FETs) have been shown to be suitable for Escherichia coli ( E. coli ) detection. Here, the G-FETs for bacterial detection are modeled and simulated with COMSOL Multiphysics to understand the operation of the biosensors. The motion of E. coli cells in electrolyte and the surface charge of graphene induced by E. coli are systematically investigated. The comparison between the simulation and experimental data proves the sensing probe size to be a key parameter affecting the surface charge of graphene induced by bacteria. Finally, the relationship among the change in source-drain current (∆ I ds ), graphene-bacteria distance and bacterial concentration is established. The shorter graphene-bacteria distance and higher bacterial concentration give rise to better sensing performance (larger ∆ I ds ) of the G-FETs biosensors. The simulation here could serve as a guideline for the design and optimization of G-FET biosensors for various applications.

Detection of Escherichia Coli Bacteria in Wastewater by using Graphene as a Sensing Material

NASA Astrophysics Data System (ADS)

Wibowo, K. M.; Sahdan, M. Z.; Ramli, N. I.; Muslihati, A.; Rosni, N.; Tsen, V. H.; Saim, H.; Ahmad, S. A.; Sari, Y.; Mansor, Z.

2018-04-01

Graphene is a family of carbon bonded in hexagonal honeycomb crystalline structure that has many superior properties. It was very suitable to be applied on sensor application due to the superior properties on electrical, physical, and optical. Furthermore, graphene also provide a large detection area since it has 2D structure. In this research, we develop graphene as a nanosensor for detection of Escherichia coli (E. coli) bacteria. The sample E. coli bacteria were cultured from domestic wastewater by using plate culture method and then isolated to get pure single colony. The serial dilution was performed to create different concentration of bacteria. Field emission scanning electron microscope and biochemical test were performed to ensure the sample genuinely target E. coli that defined by the physical size and optical properties. Raman spectroscopy measurements were also performed on the grapheme films, and it was found that the ratio of G peak and D peak intensity changing do to the presence of E. coli. The electrical properties of graphene shows the increasing number of the bacteria 4 to 273 cfu result in decreasing the resistance from 4.371 to 3.903 ohm gradually.

[The significance of biofilm for the treatment of infections in orthopedic surgery : 2017 Update].

PubMed

Scheuermann-Poley, C; Wagner, C; Hoffmann, J; Moter, A; Willy, C

2017-06-01

The increase in endoprosthetic and osteosynthetic surgical treatment is associated with a simultaneous increase in implant-associated infections (surgical site infections, SSI). Biofilms appear to play a significant role in the diagnosis and treatment of these infections and heavily contaminated wounds. This article aims to provide a current overview of biofilm and its relevance in orthopedic surgery. A computer-assisted literature search of MedLine (PubMed) was performed using key word combinations with "biofilm" (as of March 2017). Biofilm, a polymicrobial organization and life form surrounded by a polysaccharide matrix, refers to an adaptation strategy of bacteria in unfavorable living conditions (e. g. under antibiotic therapy). Biofilms can develop after 6 h in highly contaminated wounds. In acute and chronic infections, biofilms can occur in 30-80 % of the cases. Only planktonic bacteria (high metabolic activity, cultivable) can be detected in standard microbiological cultures, biofilms, however, cannot. Molecular microscopic methods, such as fluorescence in situ hybridization (FISH), enable the detection of bacteria in biofilms. The core concepts of anti-biofilm therapy include the prevention of biofilm and early surgical debridement, followed by the local and/or systemic administration of antibiotics as well as the local application of antiseptics. The development of biofilm should be anticipated in strongly contaminated wounds as well as in acute and chronic infection sites. The best strategy to combat biofilms is to prevent their development. Standard microbiological culture methods do not enable the detection of biofilm. Therefore, the implementation of molecular biological detection methods (z. B. FISH) is important. Further anti-biofilm strategies are being investigated experimentally, but there are no real options for clinical use as of yet.

Genetic markers for detection of Escherichia coli K-12 harboring ampicillin-resistance plasmid from an industrial wastewater treatment effluent pond.

PubMed

Simões, G A R; Xavier, M A S; Oliveira, D A; Menezes, E V; Magalhães, S S G; Gandra, J A C D; Xavier, A R E O

2016-06-17

Biotechnology industries that use recombinant DNA technology are potential sources for release of genetically modified organisms to the environment. Antibiotic-resistance marker genes are commonly used for recombinant bacteria selection. One example is the marker gene coding for β-lactamase (bla) in plasmids found in Escherichia coli K-12. The aim of this study was to provide an approach to develop a molecular method for genetic marker detection in E. coli K-12 harboring bla genes from an industrial wastewater treatment effluent pond (IWTEP). For the detection of bla and Achromobacter lyticus protease I (api) genes in samples from IWTEP, we employed multiplex polymerase chain reaction (PCR) using E. coli K-12 genetic marker detection primers, previously described in the literature, and primers designed in our laboratory. The microbiological screening method resulted in 22 bacterial colony-forming units isolated from three different IWTEP harvesting points. The multiplex PCR amplicons showed that five isolates were positive for the bla gene marker and negative for the E. coli K-12 and api genes. The 16S rRNA regions of positive microorganisms carrying the bla gene were genotyped by the MicroSeq®500 system. The bacteria found were Escherichia spp (3/5), Chromobacterium spp (1/5), and Aeromonas spp (1/5). None of the 22 isolated microorganisms presented the molecular pattern of E. coli K-12 harboring the bla gene. The presence of microorganisms positive for the bla gene and negative for E. coli K-12 harboring bla genes at IWTEP suggests that the ampicillin resistance found in the isolated bacteria could be from microorganisms other than the E. coli K-12 strain harboring plasmid.

Quantification of carious pathogens in the interdental microbiota of young caries-free adults

PubMed Central

Inquimbert, Camille; Tramini, Paul; Molinari, Nicolas; Carrouel, Florence

2017-01-01

Background The majority of caries lesions in adults occur on the proximal tooth surfaces of the posterior teeth. A comprehensive study of the composition of the oral microbiota is fundamental for a better understanding of the etiology of interdental caries. Methods Twenty-five caries-free subjects (20–35 years old) were enrolled in the study. The interdental biofilm of four interdental sites were collected. The real-time polymerase chain reaction (PCR) methodology were used to quantify (i) the following bacteria: Streptococcus spp., Streptococcus mutans, Lactobacillus spp., Enterococcus spp., and Enterococcus faecalis; (ii) the fungus Candida albicans; and (iii) total bacteria. Results Streptococcus spp. was the most abundant species, followed by Lactobacillus spp. and Enterococcus spp. Streptococcus spp. and Lactobacillus spp. were detected at all tested sites and Enterococcus spp. at 99% of sites. S. mutans was detected at only 28% of the tested sites and C. albicans was detected at 11% of sites. E. faecalis was never detected. In 54.5% of the biofilm inhabited by C. albicans, S. mutans was present. Moreover, 28% of the ID sites co-expressed S. mutans and Lactobacillus spp. The studied pathogens were organized into two correlated groups of species. Strikingly, the fungus C. albicans and the bacteria Enterococcus spp. cluster together, whereas Streptococcus spp., S. mutans and Lactobacillus spp. form one distinct cluster. Conclusion The interdental biofilm of young caries-free adults is comprised of pathogens that are able to induce interproximal caries. That several of these pathogens are implicated in heart disease or other systemic diseases is an argument for the disruption of interdental biofilms using daily oral hygiene. PMID:29016613

The spread of carbapenemase-producing bacteria in Africa: a systematic review

PubMed Central

Manenzhe, Rendani I.; Zar, Heather J.; Nicol, Mark P.; Kaba, Mamadou

2015-01-01

Background Carbapenems are the last line of defence against ever more prevalent MDR Gram-negative bacteria, but their efficacy is threatened worldwide by bacteria that produce carbapenemase enzymes. The epidemiology of bacteria producing carbapenemases has been described in considerable detail in Europe, North America and Asia; however, little is known about their spread and clinical relevance in Africa. Methods We systematically searched in PubMed, EBSCOhost, Web of Science, Scopus, Elsevier Masson Consulte and African Journals Online, international conference proceedings, published theses and dissertations for studies reporting on carbapenemase-producing bacteria in Africa. We included articles published in English or French up to 28 February 2014. We calculated the prevalence of carbapenemase producers only including studies where the total number of isolates tested was at least 30. Results Eighty-three studies were included and analysed. Most studies were conducted in North Africa (74%, 61/83), followed by Southern Africa (12%, 10/83), especially South Africa (90%, 9/10), West Africa (8%, 7/83) and East Africa (6%, 6/83). Carbapenemase-producing bacteria were isolated from humans, the hospital environment and community environmental water samples, but not from animals. The prevalence of carbapenemase-producing isolates in hospital settings ranged from 2.3% to 67.7% in North Africa and from 9% to 60% in sub-Saharan Africa. Conclusions Carbapenemase-producing bacteria have been described in many African countries; however, their prevalence is poorly defined and has not been systematically studied. Antibiotic stewardship and surveillance systems, including molecular detection and genotyping of resistant isolates, should be implemented to monitor and reduce the spread of carbapenemase-producing bacteria. PMID:25261423

Viable bacterial population and persistence of foodborne pathogens on the pear carpoplane.

PubMed

Duvenage, Francois J; Duvenage, Stacey; Du Plessis, Erika M; Volschenk, Quinton; Korsten, Lise

2017-03-01

Knowledge on the culturable bacteria and foodborne pathogen presence on pears is important for understanding the impact of postharvest practices on food safety assurance. Pear fruit bacteria were investigated from the point of harvest, following chlorine drenching and after controlled atmosphere (CA) storage to assess the impact on natural bacterial populations and potential foodborne pathogens. Salmonella spp. and Listeria monocytogenes were detected on freshly harvested fruit in season one. During season one, chemical drenching and CA storage did not have a significant effect on the bacterial load of orchard pears, except for two farms where the populations were lower 'after CA storage'. During season two, bacterial populations of orchard pears from three of the four farms increased significantly following drenching; however, the bacterial load decreased 'after CA storage'. Bacteria isolated following enumeration included Enterobacteriaceae, Microbacteriaceae, Pseudomonadaceae and Bacillaceae, with richness decreasing 'after drench' and 'after CA storage'. Salmonella spp. and L. monocytogenes were not detected after postharvest practices. Postharvest practices resulted in decreased bacterial species richness. Understanding how postharvest practices have an impact on the viable bacterial populations of pear fruit will contribute to the development of crop-specific management systems for food safety assurance. © 2016 Society of Chemical Industry. © 2016 Society of Chemical Industry.