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Objectives: Primary Objective: To determine the efficacy of a candidate antiviral on time to virological cure compared to standard of care within 14 days of randomisation Secondary objectives: * To determine the safety of the antiviral * To determine the clinical benefit of the antiviral over placebo according to the WHO 7-point ordinal scale * To determine the clinical benefit of the antiviral over placebo on time to resolution of clinical symptoms * To determine the effect of the antiviral over placebo on biomarkers of inflammation and immune activation Trial design: This is a multi-centre, triple-blind, randomised placebo controlled phase II, 2-arm trial with parallel-group design with allocation ratio 1:1. Participant(s): Inclusion Criteria: * Provision of informed consent by the participant * Age >=18 years * Confirmed SARS-CoV-2 by nucleic acid testing in the past 5 days * COVID-19 related symptom initiation within 5 days * Female patients of childbearing potential must have a negative pregnancy test at Screening. Female patients of childbearing potential and fertile male patients who are sexually active with a female of childbearing potential must use highly effective methods of contraception throughout the study and for 1 week following the last dose of study treatment. Exclusion criteria: * Known allergy to the study medication * Is on another clinical trial investigating an antiviral treatment for COVID-19 * Pregnancy * Patients with severe hepatic dysfunction equivalent to Grade C in the Child-Pugh classification * Patients with renal impairment requiring dialysis * Is deemed by the Investigator to be ineligible for any reason Participants will be recruited from, and the study visits will take place at Alfred Hospital, Monash Health, Austin Health in Victoria, Australia for hospitalised participants as well as recruitment in the community in participants homes for eligible people not requiring hospitalisation. Intervention and comparator: The first candid

Objectives: Primary Objective: To determine the efficacy of a candidate antiviral on time to virological cure compared to standard of care within 14 days of randomisation Secondary objectives: * To determine the safety of the antiviral * To determine the clinical benefit of the antiviral over placebo according to the WHO 7-point ordinal scale * To determine the clinical benefit of the antiviral over placebo on time to resolution of clinical symptoms * To determine the effect of the antiviral over placebo on biomarkers of inflammation and immune activation Trial design: This is a multi-centre, triple-blind, randomised placebo controlled phase II, 2-arm trial with parallel-group design with allocation ratio 1:1. Participant(s): Inclusion Criteria: * Provision of informed consent by the participant * Age >=18 years * Confirmed SARS-CoV-2 by nucleic acid testing in the past 5 days * COVID-19 related symptom initiation within 5 days * Female patients of childbearing potential must have a negative pregnancy test at Screening. Female patients of childbearing potential and fertile male patients who are sexually active with a female of childbearing potential must use highly effective methods of contraception throughout the study and for 1 week following the last dose of study treatment. Exclusion criteria: * Known allergy to the study medication * Is on another clinical trial investigating an antiviral treatment for COVID-19 * Pregnancy * Patients with severe hepatic dysfunction equivalent to Grade C in the Child-Pugh classification * Patients with renal impairment requiring dialysis * Is deemed by the Investigator to be ineligible for any reason Participants will be recruited from, and the study visits will take place at Alfred Hospital, Monash Health, Austin Health in Victoria, Australia for hospitalised participants as well as recruitment in the community in participants homes for eligible people not requiring hospitalisation. Intervention and comparator: The first candid

We present on-chip implementation of an isothermal nucleic acid amplification assay with optomagnetic detection. Isothermal rolling circle amplification (RCA) was designed for the highly specific detection of a single point mutation in the sub-sequence of the katG catalase peroxidase gene responsible for resistance against the antibiotic isoniazid in Mycobacterium tuberculosis. The assay was integrated using streptavidin coated magnetic microbeads (MMBs) as a movable substrate to transport the synthetic katG target and products between the sequential processing steps. RCA products were detected via optomagnetic measurements of the binding of functionalized magnetic nanoparticles (MNPs). Binding of MNPs to RCA products lead to an increase of their hydrodynamic size and consequently a shift of the optomagnetic signal to lower frequencies. We first analyzed the affinity of DNA probes used for target capture on MMBs and binding of MNPs to RCA products, respectively. This enabled us to choose conditions where amplification products were released from the MMBs and their binding to MNPs was optimal. The optimized assay was transferred to a polymer chip with three connected chambers filled with reaction buffers for capture, RCA and detection. The chip was operated in a setup integrating temperature control, transport of the DNA on magnetic carriers and dose-dependent detection of MNP clustering. For a total assay time of about 2 h, we demonstrated specific detection of the antibiotic resistance in synthetic Tuberculosis DNA target with a limit of detection of 4 pM.

Waterborne microbial pathogen detection via nucleic acid analysis on portable microfluidic devices is a growing area of research, development, and application. Traditional polymerase chain reaction (PCR)-based nucleic acid analysis detects total extracted DNA, but cannot differentiate live and dead cells. A propidium monoazide (PMA) pretreatment step before PCR can effectively exclude DNA from nonviable cells, as PMA can selectively diffuse through compromised cell membranes and intercalate with DNA to form DNA-PMA complex upon light exposure. The complex strongly inhibits the amplification of the bound DNA in PCR, and thus, only cells with intact cell membranes are detected. Herein, this study reports the development of a microfluidic device to carry out PMA pretreatment 'on-chip'. Chip design was guided by computer simulations, and prototypes were fabricated using a high-resolution 3D printer. The optimized design utilizes split and recombine mixers for initial PMA-sample mixing and a serpentine flow channel containing herringbone structures for dark and light incubation. On-chip PMA pretreatment to differentiate live and dead bacterial cells in buffer and natural pond water samples was successfully demonstrated.

Nucleic acids are fundamental molecules of living organisms functioning essentially as the molecular information carriers of life. From how an organism is built to how it responds to external conditions, all of it, can be found in the form of nucleic acid sequences inside every single cell of every life form on earth. Therefore, accessing these sequences provides key information regarding the molecular identity and functional state of any living organism, this is very useful for areas like biomedicine, where accessing and understanding these molecular signatures is the key to develop strategies to understand, treat and diagnose diseases. Decades of research and technological advancements have led to the development of a number of molecular tools and engineering technologies that allow accessing the information contained in the nucleic acids. This thesis provides a general overview of the tools and technologies available for nucleic acid analysis, and proposes an illustrative concept on how molecular tools and emergent technologies can be combined in a modular fashion to design methods for addressing different biomedical questions. The studies included in this thesis, are focused on the particular use of the molecular tools named: padlock and selector probes, rolling circle amplification, and fluorescence detection of single molecules in combination with microfluidics and portable microscopy. By using this combination, it became possible to design and demonstrate novel approaches for integrated nucleic acid analysis, inexpensive digital quantification, mobile-phone based diagnostics and the description of viral infections. These studies represent a step forward towards the adoption of the selected group of tools and technologies, for the design and building of methods that can be used as powerful alternatives to conventional tools used in molecular diagnostics and virology., At the time of the doctoral defense, the following paper was unpublished and had a status as follows: Paper 1: Manuscript.

Nucleic acids are fundamental molecules of living organisms functioning essentially as the molecular information carriers of life. From how an organism is built to how it responds to external conditions, all of it, can be found in the form of nucleic acid sequences inside every single cell of every life form on earth. Therefore, accessing these sequences provides key information regarding the molecular identity and functional state of any living organism, this is very useful for areas like biomedicine, where accessing and understanding these molecular signatures is the key to develop strategies to understand, treat and diagnose diseases. Decades of research and technological advancements have led to the development of a number of molecular tools and engineering technologies that allow accessing the information contained in the nucleic acids. This thesis provides a general overview of the tools and technologies available for nucleic acid analysis, and proposes an illustrative concept on how molecular tools and emergent technologies can be combined in a modular fashion to design methods for addressing different biomedical questions. The studies included in this thesis, are focused on the particular use of the molecular tools named: padlock and selector probes, rolling circle amplification, and fluorescence detection of single molecules in combination with microfluidics and portable microscopy. By using this combination, it became possible to design and demonstrate novel approaches for integrated nucleic acid analysis, inexpensive digital quantification, mobile-phone based diagnostics and the description of viral infections. These studies represent a step forward towards the adoption of the selected group of tools and technologies, for the design and building of methods that can be used as powerful alternatives to conventional tools used in molecular diagnostics and virology., At the time of the doctoral defense, the following paper was unpublished and had a status as follows: Paper 1: Manuscript.

Nucleic acids are fundamental molecules of living organisms functioning essentially as the molecular information carriers of life. From how an organism is built to how it responds to external conditions, all of it, can be found in the form of nucleic acid sequences inside every single cell of every life form on earth. Therefore, accessing these sequences provides key information regarding the molecular identity and functional state of any living organism, this is very useful for areas like biomedicine, where accessing and understanding these molecular signatures is the key to develop strategies to understand, treat and diagnose diseases. Decades of research and technological advancements have led to the development of a number of molecular tools and engineering technologies that allow accessing the information contained in the nucleic acids. This thesis provides a general overview of the tools and technologies available for nucleic acid analysis, and proposes an illustrative concept on how molecular tools and emergent technologies can be combined in a modular fashion to design methods for addressing different biomedical questions. The studies included in this thesis, are focused on the particular use of the molecular tools named: padlock and selector probes, rolling circle amplification, and fluorescence detection of single molecules in combination with microfluidics and portable microscopy. By using this combination, it became possible to design and demonstrate novel approaches for integrated nucleic acid analysis, inexpensive digital quantification, mobile-phone based diagnostics and the description of viral infections. These studies represent a step forward towards the adoption of the selected group of tools and technologies, for the design and building of methods that can be used as powerful alternatives to conventional tools used in molecular diagnostics and virology., At the time of the doctoral defense, the following paper was unpublished and had a status as follows: Paper 1: Manuscript.

Nucleic acids are fundamental molecules of living organisms functioning essentially as the molecular information carriers of life. From how an organism is built to how it responds to external conditions, all of it, can be found in the form of nucleic acid sequences inside every single cell of every life form on earth. Therefore, accessing these sequences provides key information regarding the molecular identity and functional state of any living organism, this is very useful for areas like biomedicine, where accessing and understanding these molecular signatures is the key to develop strategies to understand, treat and diagnose diseases. Decades of research and technological advancements have led to the development of a number of molecular tools and engineering technologies that allow accessing the information contained in the nucleic acids. This thesis provides a general overview of the tools and technologies available for nucleic acid analysis, and proposes an illustrative concept on how molecular tools and emergent technologies can be combined in a modular fashion to design methods for addressing different biomedical questions. The studies included in this thesis, are focused on the particular use of the molecular tools named: padlock and selector probes, rolling circle amplification, and fluorescence detection of single molecules in combination with microfluidics and portable microscopy. By using this combination, it became possible to design and demonstrate novel approaches for integrated nucleic acid analysis, inexpensive digital quantification, mobile-phone based diagnostics and the description of viral infections. These studies represent a step forward towards the adoption of the selected group of tools and technologies, for the design and building of methods that can be used as powerful alternatives to conventional tools used in molecular diagnostics and virology., At the time of the doctoral defense, the following paper was unpublished and had a status as follows: Paper 1: Manuscript.

Nucleic acids are fundamental molecules of living organisms functioning essentially as the molecular information carriers of life. From how an organism is built to how it responds to external conditions, all of it, can be found in the form of nucleic acid sequences inside every single cell of every life form on earth. Therefore, accessing these sequences provides key information regarding the molecular identity and functional state of any living organism, this is very useful for areas like biomedicine, where accessing and understanding these molecular signatures is the key to develop strategies to understand, treat and diagnose diseases. Decades of research and technological advancements have led to the development of a number of molecular tools and engineering technologies that allow accessing the information contained in the nucleic acids. This thesis provides a general overview of the tools and technologies available for nucleic acid analysis, and proposes an illustrative concept on how molecular tools and emergent technologies can be combined in a modular fashion to design methods for addressing different biomedical questions. The studies included in this thesis, are focused on the particular use of the molecular tools named: padlock and selector probes, rolling circle amplification, and fluorescence detection of single molecules in combination with microfluidics and portable microscopy. By using this combination, it became possible to design and demonstrate novel approaches for integrated nucleic acid analysis, inexpensive digital quantification, mobile-phone based diagnostics and the description of viral infections. These studies represent a step forward towards the adoption of the selected group of tools and technologies, for the design and building of methods that can be used as powerful alternatives to conventional tools used in molecular diagnostics and virology., At the time of the doctoral defense, the following paper was unpublished and had a status as follows: Paper 1: Manuscript.

Nucleic acids are fundamental molecules of living organisms functioning essentially as the molecular information carriers of life. From how an organism is built to how it responds to external conditions, all of it, can be found in the form of nucleic acid sequences inside every single cell of every life form on earth. Therefore, accessing these sequences provides key information regarding the molecular identity and functional state of any living organism, this is very useful for areas like biomedicine, where accessing and understanding these molecular signatures is the key to develop strategies to understand, treat and diagnose diseases. Decades of research and technological advancements have led to the development of a number of molecular tools and engineering technologies that allow accessing the information contained in the nucleic acids. This thesis provides a general overview of the tools and technologies available for nucleic acid analysis, and proposes an illustrative concept on how molecular tools and emergent technologies can be combined in a modular fashion to design methods for addressing different biomedical questions. The studies included in this thesis, are focused on the particular use of the molecular tools named: padlock and selector probes, rolling circle amplification, and fluorescence detection of single molecules in combination with microfluidics and portable microscopy. By using this combination, it became possible to design and demonstrate novel approaches for integrated nucleic acid analysis, inexpensive digital quantification, mobile-phone based diagnostics and the description of viral infections. These studies represent a step forward towards the adoption of the selected group of tools and technologies, for the design and building of methods that can be used as powerful alternatives to conventional tools used in molecular diagnostics and virology., At the time of the doctoral defense, the following paper was unpublished and had a status as follows: Paper 1: Manuscript.

Rapid and timely diagnosis of infectious diseases is a critical determinant of clinical outcomes and general public health. For the detection of various pathogens, microfluidics-based platforms offer many advantages, including speed, cost, portability, high throughput, and automation. This review provides an overview of the recent advances in microfluidic technologies for point-of-care (POC) diagnostics for infectious diseases. The key aspects of such technologies for the development of a fully integrated POC platform are introduced, including sample preparation, on-chip nucleic acid analysis and immunoassay, and system integration/automation. The current challenges to practical implementation of this technology are discussed together with future perspectives. (C) 2014 Elsevier B.V. All rights reserved.

We report a case of Acanthamoeba encephalitis diagnosed from an antemortem brain biopsy specimen, where the organism was first isolated in mycobacterial liquid medium and first identified by using a sequence generated by a commercial panfungal sequencing assay. We correlate susceptibility results with clinical outcome.Copyright © 2015, American Society for Microbiology. All Rights Reserved.

We report a case of Acanthamoeba encephalitis diagnosed from an antemortem brain biopsy specimen, where the organism was first isolated in mycobacterial liquid medium and first identified by using a sequence generated by a commercial panfungal sequencing assay. We correlate susceptibility results with clinical outcome.Copyright © 2015, American Society for Microbiology. All Rights Reserved.

Rapid and timely diagnosis of infectious diseases is a critical determinant of clinical outcomes and general public health. For the detection of various pathogens, microfluidics-based platforms offer many advantages, including speed, cost, portability, high throughput, and automation. This review provides an overview of the recent advances in microfluidic technologies for point-of-care (POC) diagnostics for infectious diseases. The key aspects of such technologies for the development of a fully integrated POC platform are introduced, including sample preparation, on-chip nucleic acid analysis and immunoassay, and system integration/automation. The current challenges to practical implementation of this technology are discussed together with future perspectives. (C) 2014 Elsevier B.V. All rights reserved.

Clinical laboratories are increasingly using molecular tests for methicillin-resistant Staphylococcus aureus (MRSA) screening. However, primers have to be targeted to a variable chromosomal region, the staphylococcal cassette chromosome mec (SCCmec). We initially screened 726 MRSA isolates from a single UK hospital trust by recombinase polymerase amplification (RPA), a novel, isothermal alternative to PCR. Undetected isolates were further characterised using multilocus sequence, spa typing and whole genome sequencing. 96% of our tested phenotypically MRSA isolates contained one of the six orfX-SCCmec junctions our RPA test and commercially available molecular tests target. However 30 isolates could not be detected. Sequencing of 24 of these isolates demonstrated recombinations within the SCCmec element with novel insertions that interfered with the RPA, preventing identification as MRSA. This result suggests that clinical laboratories cannot rely solely upon molecular assays to reliably detect all methicillin-resistance. The presence of significant recombinations in the SCCmec element, where the majority of assays target their primers, suggests that there will continue to be isolates that escape identification. We caution that dependence on amplification-based molecular assays will continue to result in failure to diagnose a small proportion (?4%) of MRSA isolates, unless the true level of SCCmec natural diversity is determined by whole genome sequencing of a large collection of MRSA isolates. © 2014 Hill-Cawthorne et al.

The recent technological advances in micro/nanotechnology present new opportunities to combine microfluidics with microarray technology for the development of small, sensitive, single-use, point-of-care molecular diagnostic devices. As such, the integration of microarray and plastic microfluidic systems is an attractive low-cost alternative to glass based microarray systems. This paper presents the integration of a DNA microarray and an all-polymer microfluidic foil system with integrated thin film heaters, which demonstrate DNA analysis based on melting curve analysis (MCA). A novel micro-heater concept using semi-transparent copper heaters manufactured by roll-to-roll and lift-off on polyethylene naphthalate (PEN) foil has been developed. Using a mesh structure, heater surfaces have been realized in only one single metallization step, providing more efficient and homogenous heating characteristics than conventional meander heaters. A robust DNA microarray spotting protocol was adapted on Parylene C coated heater-foils, using co-polymer poly(DMA-NAS-MAPS) to enable covalent immobilization of DNA. The heaters were integrated in a microfluidic channel using lamination foils and MCA of the spotted DNA duplexes showed single based discrimination of mismatched over matched target DNA-probes. Finally, as a proof of principle, we perform MCA on PCR products to detect the Leu7Pro polymorphism of the neutropeptide Y related to increased risk of Type II diabetes, BMI and depression., QC 20130614

The recent technological advances in micro/nanotechnology present new opportunities to combine microfluidics with microarray technology for the development of small, sensitive, single-use, point-of-care molecular diagnostic devices. As such, the integration of microarray and plastic microfluidic systems is an attractive low-cost alternative to glass based microarray systems. This paper presents the integration of a DNA microarray and an all-polymer microfluidic foil system with integrated thin film heaters, which demonstrate DNA analysis based on melting curve analysis (MCA). A novel micro-heater concept using semi-transparent copper heaters manufactured by roll-to-roll and lift-off on polyethylene naphthalate (PEN) foil has been developed. Using a mesh structure, heater surfaces have been realized in only one single metallization step, providing more efficient and homogenous heating characteristics than conventional meander heaters. A robust DNA microarray spotting protocol was adapted on Parylene C coated heater-foils, using co-polymer poly(DMA-NAS-MAPS) to enable covalent immobilization of DNA. The heaters were integrated in a microfluidic channel using lamination foils and MCA of the spotted DNA duplexes showed single based discrimination of mismatched over matched target DNA-probes. Finally, as a proof of principle, we perform MCA on PCR products to detect the Leu7Pro polymorphism of the neutropeptide Y related to increased risk of Type II diabetes, BMI and depression., QC 20130614

The recent technological advances in micro/nanotechnology present new opportunities to combine microfluidics with microarray technology for the development of small, sensitive, single-use, point-of-care molecular diagnostic devices. As such, the integration of microarray and plastic microfluidic systems is an attractive low-cost alternative to glass based microarray systems. This paper presents the integration of a DNA microarray and an all-polymer microfluidic foil system with integrated thin film heaters, which demonstrate DNA analysis based on melting curve analysis (MCA). A novel micro-heater concept using semi-transparent copper heaters manufactured by roll-to-roll and lift-off on polyethylene naphthalate (PEN) foil has been developed. Using a mesh structure, heater surfaces have been realized in only one single metallization step, providing more efficient and homogenous heating characteristics than conventional meander heaters. A robust DNA microarray spotting protocol was adapted on Parylene C coated heater-foils, using co-polymer poly(DMA-NAS-MAPS) to enable covalent immobilization of DNA. The heaters were integrated in a microfluidic channel using lamination foils and MCA of the spotted DNA duplexes showed single based discrimination of mismatched over matched target DNA-probes. Finally, as a proof of principle, we perform MCA on PCR products to detect the Leu7Pro polymorphism of the neutropeptide Y related to increased risk of Type II diabetes, BMI and depression., QC 20130614

The recent technological advances in micro/nanotechnology present new opportunities to combine microfluidics with microarray technology for the development of small, sensitive, single-use, point-of-care molecular diagnostic devices. As such, the integration of microarray and plastic microfluidic systems is an attractive low-cost alternative to glass based microarray systems. This paper presents the integration of a DNA microarray and an all-polymer microfluidic foil system with integrated thin film heaters, which demonstrate DNA analysis based on melting curve analysis (MCA). A novel micro-heater concept using semi-transparent copper heaters manufactured by roll-to-roll and lift-off on polyethylene naphthalate (PEN) foil has been developed. Using a mesh structure, heater surfaces have been realized in only one single metallization step, providing more efficient and homogenous heating characteristics than conventional meander heaters. A robust DNA microarray spotting protocol was adapted on Parylene C coated heater-foils, using co-polymer poly(DMA-NAS-MAPS) to enable covalent immobilization of DNA. The heaters were integrated in a microfluidic channel using lamination foils and MCA of the spotted DNA duplexes showed single based discrimination of mismatched over matched target DNA-probes. Finally, as a proof of principle, we perform MCA on PCR products to detect the Leu7Pro polymorphism of the neutropeptide Y related to increased risk of Type II diabetes, BMI and depression., QC 20130614

The recent technological advances in micro/nanotechnology present new opportunities to combine microfluidics with microarray technology for the development of small, sensitive, single-use, point-of-care molecular diagnostic devices. As such, the integration of microarray and plastic microfluidic systems is an attractive low-cost alternative to glass based microarray systems. This paper presents the integration of a DNA microarray and an all-polymer microfluidic foil system with integrated thin film heaters, which demonstrate DNA analysis based on melting curve analysis (MCA). A novel micro-heater concept using semi-transparent copper heaters manufactured by roll-to-roll and lift-off on polyethylene naphthalate (PEN) foil has been developed. Using a mesh structure, heater surfaces have been realized in only one single metallization step, providing more efficient and homogenous heating characteristics than conventional meander heaters. A robust DNA microarray spotting protocol was adapted on Parylene C coated heater-foils, using co-polymer poly(DMA-NAS-MAPS) to enable covalent immobilization of DNA. The heaters were integrated in a microfluidic channel using lamination foils and MCA of the spotted DNA duplexes showed single based discrimination of mismatched over matched target DNA-probes. Finally, as a proof of principle, we perform MCA on PCR products to detect the Leu7Pro polymorphism of the neutropeptide Y related to increased risk of Type II diabetes, BMI and depression., QC 20130614

Epithelial-neutrophil activating peptide-78 (ENA-78) induces neutrophil migration, an early response to viral infection. Rhinovirus serotype 16 (RV16) was used to infect primary bronchial epithelial cells and a cell line (BEAS-2B). Release of ENA-78 protein was measured by enzyme-linked immunosorbent assay, ENA-78 mRNA production was quantified by reverse-transcription polymerase chain reaction, and ENA-78 promoter activity was assessed by use of a promoter construct. After infection with RV16, ENA-78 protein and mRNA increased significantly, and RV16 induced 3-fold increases in ENA-78 gene transcription. Nasal ENA-78 measured in patients with asthma with and without RV infection was more elevated in patients with RV infection present. Our study demonstrates that ENA-78 is produced in bronchial epithelial cells in response to RV16 infection. With other chemokines, it may be an important initiator of neutrophil airway inflammation during RV common colds and thus may play a role in the development of virus-associated airway pathologies.

Background and objective: National surveillance of invasive pneumococcal disease (IPD) includes serotyping Streptococcus pneumoniae (SP) isolates from sterile site cultures. PCR is more sensitive and can identify more SP serotypes (STs) in culture-negative samples. The aim of this study was to determine whether enhanced surveillance of childhood empyema, using PCR, provides additional serotype information compared with conventional surveillance. Method(s): Pleural fluid (PF) from children with empyema were cultured and tested by PCR to identify SP, targeting the autolysin gene (lytA). Multiplex PCR-based reverse line blot assay was used to identify SP STs. Corresponding IPD surveillance and serotype data were obtained from the National Notifiable Diseases Surveillance System (NNDSS). Result(s): Eighty-nine children with empyema, aged <=16 years, were recruited between April 2008 and March 2009, inclusive. SP was isolated from 5/84 (5.9%) PF cultures and by PCR in 43/79 (54.4%) PF samples. © 2011 Asian Pacific Society of Respirology.

Epithelial-neutrophil activating peptide-78 (ENA-78) induces neutrophil migration, an early response to viral infection. Rhinovirus serotype 16 (RV16) was used to infect primary bronchial epithelial cells and a cell line (BEAS-2B). Release of ENA-78 protein was measured by enzyme-linked immunosorbent assay, ENA-78 mRNA production was quantified by reverse-transcription polymerase chain reaction, and ENA-78 promoter activity was assessed by use of a promoter construct. After infection with RV16, ENA-78 protein and mRNA increased significantly, and RV16 induced 3-fold increases in ENA-78 gene transcription. Nasal ENA-78 measured in patients with asthma with and without RV infection was more elevated in patients with RV infection present. Our study demonstrates that ENA-78 is produced in bronchial epithelial cells in response to RV16 infection. With other chemokines, it may be an important initiator of neutrophil airway inflammation during RV common colds and thus may play a role in the development of virus-associated airway pathologies.

Background and objective: National surveillance of invasive pneumococcal disease (IPD) includes serotyping Streptococcus pneumoniae (SP) isolates from sterile site cultures. PCR is more sensitive and can identify more SP serotypes (STs) in culture-negative samples. The aim of this study was to determine whether enhanced surveillance of childhood empyema, using PCR, provides additional serotype information compared with conventional surveillance. Method(s): Pleural fluid (PF) from children with empyema were cultured and tested by PCR to identify SP, targeting the autolysin gene (lytA). Multiplex PCR-based reverse line blot assay was used to identify SP STs. Corresponding IPD surveillance and serotype data were obtained from the National Notifiable Diseases Surveillance System (NNDSS). Result(s): Eighty-nine children with empyema, aged <=16 years, were recruited between April 2008 and March 2009, inclusive. SP was isolated from 5/84 (5.9%) PF cultures and by PCR in 43/79 (54.4%) PF samples. © 2011 Asian Pacific Society of Respirology.

Real-time polymerase chain reaction (PCR) is the current method of choice for detection and quantification of nucleic acids, especially for molecular diagnostics. Complementarity between primers and template is often crucial for PCR applications, as mismatches can severely reduce priming efficiency. However, little quantitative data on the effect of these mismatches is available. We quantitatively investigated the effects of primer-template mismatches within the 3′-end primer region on real-time PCR using the 5′-nuclease assay. Our results show that single mismatches i

This article is free to read on the publishers website A moderately thermophilic, facultatively anaerobic, amylolytic bacterium was isolated from palm wine, a tropical alcoholic beverage that was sampled in Senegal. The cells were gram positive, catalase positive, non-spore forming, rod shaped, and slightly motile with peritrichous flagella. The strain which we examined did not possess cytochrome and produced L-(+)-lactate, acetate, ethanol, and formate but not hydrogen during carbohydrate fermentation. Growth occurred at pH values ranging from 5.4 to 8.5, and optimum growth occurred at around pH 7.0. The optimum temperature for growth was around 50°C, and the upper temperature limit for growth was 58°C. The guanine-plus-cytosine content of the DNA was 38.8 ± 0.2 mol%. A sequence analysis of the 16S rRNA gene revealed that the new organism is closely related phylogenetically to members of genus Bacillus. Despite the lack of spores, we propose that on the basis of phylogenetic characteristics, the new isolate should be classified as a new Bacillus species, Bacillus thermoamylovorans. The type strain is strain DKP (= Collection of Institut Pasteur CNCM 1-1378).