scholarly journals Controlled Human Infection With Bordetella pertussis Induces Asymptomatic, Immunizing Colonization

2019 ◽  
Vol 71 (2) ◽  
pp. 403-411 ◽  
Author(s):  
Hans de Graaf ◽  
Muktar Ibrahim ◽  
Alison R Hill ◽  
Diane Gbesemete ◽  
Andrew T Vaughan ◽  
...  
Keyword(s):  
Bordetella Pertussis ◽  
Quantitative Polymerase Chain Reaction ◽  
Human Infection ◽  
Wild Type ◽  
Immunological Responses ◽  
Inoculum Dose ◽  
Colony Forming Units ◽  
Community Transmission ◽  
Polymerase Chain ◽  
Immunoglobin G

Abstract Background Bordetella pertussis is among the leading causes of vaccine-preventable deaths and morbidity globally. Human asymptomatic carriage as a reservoir for community transmission of infections might be a target of future vaccine strategies, but has not been demonstrated. Our objective was to demonstrate that asymptomatic nasopharyngeal carriage of Bordetella pertussis is inducible in humans and to define the microbiological and immunological features of presymptomatic infection. Methods Healthy subjects aged 18–45 years with an antipertussis toxin immunoglobin G (IgG) concentration of <20 international units/ml were inoculated intranasally with nonattenuated, wild-type Bordetella pertussis strain B1917. Safety, colonization, and shedding were monitored over 17 days in an inpatient facility. Colonization was assessed by culture and quantitative polymerase chain reaction. Azithromycin was administered from Day 14. The inoculum dose was escalated, aiming to colonize at least 70% of participants. Immunological responses were measured. Results There were 34 participants challenged, in groups of 4 or 5. The dose was gradually escalated from 103 colony-forming units (0% colonized) to 105 colony-forming units (80% colonized). Minor symptoms were reported in a minority of participants. Azithromycin eradicated colonization in 48 hours in 88% of colonized individuals. Antipertussis toxin IgG seroconversion occurred in 9 out of 19 colonized participants and in none of the participants who were not colonized. Nasal wash was a more sensitive method to detect colonization than pernasal swabs. No shedding of Bordetella pertussis was detected in systematically collected environmental samples. Conclusions Bordetella pertussis colonization can be deliberately induced and leads to a systemic immune response without causing pertussis symptoms. Clinical Trials Registration NCT03751514.

scholarly journals Activation of C-Type Lectin Receptor and (RIG)-I-Like Receptors Contributes to Proinflammatory Response in Middle East Respiratory Syndrome Coronavirus-Infected Macrophages

2019 ◽  
Author(s):  
Xiaoyu Zhao ◽  
Hin Chu ◽  
Bosco Ho-Yin Wong ◽  
Man Chun Chiu ◽  
Dong Wang ◽  
...  
Keyword(s):  
Middle East ◽  
Quantitative Polymerase Chain Reaction ◽  
Severe Disease ◽  
Human Infection ◽  
Middle East Respiratory Syndrome ◽  
Proinflammatory Response ◽  
Pharmacological Inhibition ◽  
Lectin Receptor ◽  
Polymerase Chain

Abstract Background Human infection with Middle East respiratory syndrome coronavirus (MERS-CoV) poses an ongoing threat to public health worldwide. The studies of MERS patients with severe disease and experimentally infected animals showed that robust viral replication and intensive proinflammatory response in lung tissues contribute to high pathogenicity of MERS-CoV. We sought to identify pattern recognition receptor (PRR) signaling pathway(s) that mediates the inflammatory cascade in human macrophages upon MERS-CoV infection. Methods The potential signaling pathways were manipulated individually by pharmacological inhibition, small interfering ribonucleic acid (siRNA) depletion, and antibody blocking. The MERS-CoV-induced proinflammatory response was evaluated by measuring the expression levels of key cytokines and/or chemokines. Reverse transcription-quantitative polymerase chain reaction assay, flow cytometry analysis, and Western blotting were applied to evaluate the activation of related PRRs and engagement of adaptors. Results MERS-CoV replication significantly upregulated C-type lectin receptor (CLR) macrophage-inducible Ca2+-dependent lectin receptor (Mincle). The role of Mincle for MERS-CoV-triggered cytokine/chemokine induction was established based on the results of antibody blockage, siRNA depletion of Mincle and its adaptor spleen tyrosine kinase (Syk), and Syk pharmacological inhibition. The cytokine and/or chemokine induction was significantly attenuated by siRNA depletion of retinoic acid-inducible-I-like receptors (RLR) or adaptor, indicating that RLR signaling also contributed to MERS-CoV-induced proinflammatory response. Conclusions The CLR and RLR pathways are activated and contribute to the proinflammatory response in MERS-CoV-infected macrophages.

Clonal analysis of deletions on chromosome 20q and JAK2-V617F in MPD suggests that del20q acts independently and is not one of the predisposing mutations for JAK2-V617F

Blood ◽  
2009 ◽  
Vol 113 (9) ◽  
pp. 2022-2027 ◽  
Author(s):  
Franz X. Schaub ◽  
Roland Jäger ◽  
Renate Looser ◽  
Hui Hao-Shen ◽  
Sylvie Hermouet ◽  
...  
Keyword(s):  
Temporal Order ◽  
Jak2 V617f ◽  
Myeloproliferative Disorders ◽  
Genetic Alterations ◽  
Parental Origin ◽  
Wild Type ◽  
Chain Reaction ◽  
Colony Forming Units ◽  
Opposite Order ◽  
Polymerase Chain

We developed a real-time copy number polymerase chain reaction assay for deletions on chromosome 20q (del20q), screened peripheral blood granulocytes from 664 patients with myeloproliferative disorders, and identified 19 patients with del20q (2.9%), of which 14 (74%) were also positive for JAK2-V617F. To examine the temporal relationship between the occurrence of del20q and JAK2-V617F, we performed colony assays in methylcellulose, picked individual burst-forming units–erythroid (BFU-E) and colony-forming units–granulocyte (CFU-G) colonies, and genotyped each colony individually for del20q and JAK2-V617F. In 2 of 9 patients, we found that some colonies with del20q carried only wild-type JAK2, whereas other del20q colonies were JAK2-V617F positive, indicating that del20q occurred before the acquisition of JAK2-V617F. However, in colonies from 3 of 9 patients, we observed the opposite order of events. The lack of a strict temporal order of occurrence makes it doubtful that del20q represents a predisposing event for JAK2-V617F. In 2 patients with JAK2-V617F and 1 patient with MPL-W515L, microsatellite analysis revealed that del20q affected chromosomes of different parental origin and/or 9pLOH occurred at least twice. The fact that rare somatic events, such as del20q or 9pLOH, occurred more than once in subclones from the same patients suggests that the myeloproliferative disorder clone carries a predisposition to acquiring such genetic alterations.

scholarly journals Redox-Dependent Structural Modification of Nucleoredoxin Triggers Defense Responses against Alternaria brassicicola in Arabidopsis

2020 ◽  
Vol 21 (23) ◽  
pp. 9196
Author(s):  
Chang Ho Kang ◽  
Joung Hun Park ◽  
Eun Seon Lee ◽  
Seol Ki Paeng ◽  
Ho Byoung Chae ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Structural Modification ◽  
Quantitative Polymerase Chain Reaction ◽  
Defense Responses ◽  
Alternaria Brassicicola ◽  
Low Molecular Weight ◽  
Biological Processes ◽  
Wild Type ◽  
Molecular Sensor ◽  
Polymerase Chain

In plants, thioredoxin (TRX) family proteins participate in various biological processes by regulating the oxidative stress response. However, their role in phytohormone signaling remains largely unknown. In this study, we investigated the functions of TRX proteins in Arabidopsis thaliana. Quantitative polymerase chain reaction (qPCR) experiments revealed that the expression of ARABIDOPSIS NUCLEOREDOXIN 1 (AtNRX1) is specifically induced by the application of jasmonic acid (JA) and upon inoculation with a necrotrophic fungal pathogen, Alternaria brassicicola. The AtNRX1 protein usually exists as a low molecular weight (LMW) monomer and functions as a reductase, but under oxidative stress AtNRX1 transforms into polymeric forms. However, the AtNRX1M3 mutant protein, harboring four cysteine-to-serine substitutions in the TRX domain, did not show structural modification under oxidative stress. The Arabidopsisatnrx1 null mutant showed greater resistance to A. brassicicola than wild-type plants. In addition, plants overexpressing both AtNRX1 and AtNRX1M3 were susceptible to A. brassicicola infection. Together, these findings suggest that AtNRX1 normally suppresses the expression of defense-responsive genes, as if it were a safety pin, but functions as a molecular sensor through its redox-dependent structural modification to induce disease resistance in plants.

scholarly journals Inoculation of fowlpox viruses coexpressing avian influenza H5 and chicken IL-15 cytokine gene stimulates diverse host immune responses

2019 ◽  
pp. 84-94 ◽  
Author(s):  
Abdul Razak Mariatulqabtiah ◽  
Nadzreeq Nor Majid ◽  
Efstathios S. Giotis ◽  
Abdul Rahman Omar ◽  
Michael A. Skinner
Keyword(s):  
Avian Influenza ◽  
Immune Responses ◽  
Quantitative Polymerase Chain Reaction ◽  
Fold Change ◽  
Post Inoculation ◽  
Immunological Responses ◽  
Host Immune Responses ◽  
Genes Expression ◽  
Specific Pathogen ◽  
Polymerase Chain

Fowlpox virus (FWPV) has been used as a recombinant vaccine vector to express antigens from several important avian pathogens. Attempts have been made to improve vaccine strains induced-host immune responses by coexpressing cytokines. This study describes the construction of recombinant FWPV (rFWPV) strain FP9 and immunological responses in specific-pathogen-free (SPF) chickens, co-expressing avian influenza virus (AIV) H5 of A/Chicken/Malaysia/5858/2004, and chicken IL-15 cytokine genes. Expression of H5 (50 kD) was confirmed by western blotting. Anti-H5 antibodies, which were measured by the haemagglutinin inhibition test, were at the highest levels at Week 3 post-inoculation in both rFWPV/H5- and rFWPV/H5/IL-15-vaccinated chickens, but decreased to undetectable levels from Week 5 onwards. CD3+/CD4+ or CD3+/CD8+T cell populations, assessed using flow cytometry, were significantly increased in both WT FP9- and rFWPV/H5-vaccinated chickens and were also higher than in rFWPV/H5/IL-15- vaccinated chickens, at Week 2. Gene expression analysis using real time quantitative polymerase chain reaction (qPCR) demonstrated upregulation of IL-15 expression in all vaccinated groups with rFWPV/H5/IL-15 having the highest fold change, at day 2 (117±51.53). Despite showing upregulation, fold change values of the IL-18 expression were below 1.00 for all vaccinated groups at day 2, 4 and 6. This study shows successful construction of rFWPV/H5 co-expressing IL-15, with modified immunogenicity upon inoculation into SPF chickens.

scholarly journals COVID-19 Community Transmission and Super Spreaders in Rural Villages from Manabi Province in the Coastal Region of Ecuador Assessed by Massive Testing of Community-Dwelling Population

2021 ◽  
Author(s):  
Maria Belén Rodriguez-Paredes ◽  
Paolo Alexander Vallejo-Janeta ◽  
Diana Morales-Jadan ◽  
Byron Freire-Paspuel ◽  
Esteban Ortiz-Prado ◽  
...  
Keyword(s):  
Rural Communities ◽  
Quantitative Polymerase Chain Reaction ◽  
Coastal Region ◽  
Community Dwelling ◽  
First Semester ◽  
Rural Villages ◽  
Viral Loads ◽  
High Prevalence ◽  
Community Transmission ◽  
Polymerase Chain

Neglected rural communities in Latin America are highly vulnerable to COVID-19 due to a poor health infrastructure and limited access to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) diagnosis. Manabí is a province of the Coastal Region of Ecuador characterized by a high prevalence of rural population living under poverty conditions. In the current study, we present the retrospective analysis of the results of a massive SARS-CoV-2 testing operation in nonhospitalized populations from Manabí carried out from August to September 2020. A total of 4,003 people from 15 cantons were tested for SARS-CoV-2 by reverse-transcriptase quantitative polymerase chain reaction, resulting in an overall infection rate of 16.13% for SARS-CoV-2, with several communities > 30%. Moreover, 29 SARS-CoV-2 super-spreader community-dwelling individuals with viral loads above 108 copies/mL were found. These results support that uncontrolled COVID-19 community transmission was happening in Manabí during the first semester of COVID-19 pandemic. This report endorses the utility of massive SARS-CoV-2 testing among asymptomatic population for control and surveillance of COVID-19.

scholarly journals Novel Characterization of Antioxidant Enzyme, 3-Mercaptopyruvate Sulfurtransferase-Knockout Mice: Overexpression of the Evolutionarily-Related Enzyme Rhodanese

Antioxidants ◽  
2019 ◽  
Vol 8 (5) ◽  
pp. 116 ◽  
Author(s):  
Noriyuki Nagahara ◽  
Mio Tanaka ◽  
Yukichi Tanaka ◽  
Takaaki Ito
Keyword(s):  
Antioxidant Enzyme ◽  
Quantitative Polymerase Chain Reaction ◽  
Polymerase Chain Reaction Analysis ◽  
Subcellular Fractionation ◽  
Liver Mitochondria ◽  
Wild Type ◽  
Mercaptopyruvate Sulfurtransferase ◽  
Related Enzyme ◽  
Polymerase Chain

The antioxidant enzyme, 3-mercaptopyruvate sulfurtransferase (MST, EC 2.8.1.2) is localized in the cytosol and mitochondria, while the evolutionarily-related enzyme, rhodanese (thiosulfate sulfurtransferase, TST, EC 2.8.1.1) is localized in the mitochondria. Recently, both enzymes have been shown to produce hydrogen sulfide and polysulfide. Subcellular fractionation of liver mitochondria revealed that the TST activity ratio of MST-knockout (KO)/wild-type mice was approximately 2.5; MST activity was detected only in wild-type mice, as expected. The ratio of TST mRNA expression of KO/wild-type mice, as measured by real-time quantitative polymerase chain reaction analysis, was approximately 3.3. It is concluded that TST is overexpressed in MST-KO mice.

Association of polymorphisms in MALAT1 with the risk of endometriosis in Southern Chinese women

2019 ◽  
Vol 102 (4) ◽  
pp. 943-949
Author(s):  
Guange Chen ◽  
Mingyao Zhang ◽  
Zongwen Liang ◽  
Sailing Chen ◽  
Feng Chen ◽  
...  
Keyword(s):  
Chinese Women ◽  
Quantitative Polymerase Chain Reaction ◽  
Uterine Cavity ◽  
Figo Stage ◽  
Wild Type ◽  
Southern Chinese ◽  
Gynecology And Obstetrics ◽  
Polymerase Chain ◽  
Unconditional Logistic Regression Model ◽  
Southern Chinese Women

Abstract Endometriosis is a common estrogen-dependent inflammatory disease characterized by the presence of endometrial-like tissue outside the uterine cavity, which causes infertility and pelvic pain. Polymorphisms in MALAT1 have been demonstrated to play crucial roles in many diseases. However, the roles of MALAT1 polymorphisms in the etiology of endometriosis have not been well documented. We genotyped three MALAT1 polymorphisms in 555 endometriosis patients and 535 female control participants using quantitative polymerase chain reaction with TaqMan probes. To estimate the associations between MALAT1 polymorphisms and endometriosis risk, an unconditional logistic regression model was conducted to calculate an odds ratio (OR) and the 95% confidence interval (CI), adjusting for age, abortion history, number of deliveries, Body Mass Index (BMI), and The International Federation of Gynecology and Obstetrics (FIGO) stage. We found that the MALAT1 rs591291 C > T polymorphism significantly enhanced endometriosis risk (heterogeneous: adjusted OR = 1.36, 95% CI = 1.00–1.85, P = 0.050; homogenous: adjusted OR = 1.55, 95% CI = 1.03–2.33, P = 0.037; dominant: adjusted OR = 1.41, 95% CI = 1.05–1.88, P = 0.021). In stratification analyses, these associations were more predominant in the patients younger than 35 years old, with a relatively high number of deliveries and with a BMI between 25 and 29.9. Compared with wild-type CCG haplotype carriers, individuals with TCC haplotypes had a higher risk of developing endometriosis. The MALAT1 rs591291 C > T polymorphism was associated with a significant increase in endometriosis risk.

AN IMPROVED METHOD FOR INHIBITOR FREE NUCLEIC ACIDS FROM POLYPHENOL RICH LEAVES OF MUSA SPP

2017 ◽  
Vol 23 (1) ◽  
Author(s):  
N.NANDHA KUMAR ◽  
K. SOURIANATHA SUNDARAM ◽  
D. SUDHAKAR ◽  
K.K. KUMAR
Keyword(s):  
Polymerase Chain Reaction ◽  
Quantitative Polymerase Chain Reaction ◽  
Proteinase K ◽  
Chain Reaction ◽  
Banana Plant ◽  
High Molecular Weight Dna ◽  
Musa Spp ◽  
Leaf Tissues ◽  
Polymerase Chain

Excessive presence of polysaccharides, polyphenol and secondary metabolites in banana plant affects the quality of DNA and it leads to difficult in isolating good quality of DNA. An optimized modified CTAB protocol for the isolation of high quality and quantity of DNA obtained from banana leaf tissues has been developed. In this protocol a slight increased salt (NaCl) concentration (2.0M) was used in the extraction buffer. Polyvinylpyrrolidone (PVP) and Octanol were used for the removal of polyphenols and polymerase chain reaction (PCR) inhibitors. Proteins like various enzymes were degraded by Proteinase K and removed by centrifugation from plant extract during the isolation process resulting in pure genomic DNA, ready to use in downstream applications including PCR, quantitative polymerase chain reaction (qPCR), ligation, restriction and sequencing. This protocol yielded a high molecular weight DNA isolated from polyphenols rich leaves of Musa spp which was free from contamination and colour. The average yields of total DNA from leaf ranged from 917.4 to 1860.9 ng/ìL. This modified CTAB protocol reported here is less time consuming 4-5h, reproducible and can be used for a broad spectrum of plant species which have polyphenol and polysaccharide compounds.

Pim-1 is up-regulated by constitutively activated FLT3 and plays a role in FLT3-mediated cell survival

Blood ◽  
2005 ◽  
Vol 105 (4) ◽  
pp. 1759-1767 ◽  
Author(s):  
Kyu-Tae Kim ◽  
Kristin Baird ◽  
Joon-Young Ahn ◽  
Paul Meltzer ◽  
Michael Lilly ◽  
...  
Keyword(s):  
Tyrosine Kinase ◽  
Quantitative Polymerase Chain Reaction ◽  
Colony Growth ◽  
Dominant Negative ◽  
Internal Tandem Duplication ◽  
Downstream Target ◽  
Before And After ◽  
Flt3 Expression ◽  
Polymerase Chain ◽  
Expression Microarrays

AbstractConstitutively activating internal tandem duplication (ITD) mutations of the receptor tyrosine kinase FLT3 (Fms-like tyrosine kinase 3) play an important role in leukemogenesis, and their presence is associated with poor prognosis in acute myeloid leukemia (AML). To better understand FLT3 signaling in leukemogenesis, we have examined the changes in gene expression induced by FLT3/ITD or constitutively activated wild-type FLT3 expression. Microarrays were used with RNA harvested before and after inhibition of FLT3 signaling. Pim-1 was found to be one of the most significantly down-regulated genes upon FLT3 inhibition. Pim-1 is a proto-oncogene and is known to be up-regulated by signal transducer and activator of transcription 5 (STAT5), which itself is a downstream target of FLT3 signaling. Quantitative polymerase chain reaction (QPCR) confirmed the microarray results and demonstrated approximately 10-fold decreases in Pim-1 expression in response to FLT3 inhibition. Pim-1 protein also decreased rapidly in parallel with decreasing autophosphorylation activity of FLT3. Enforced expression of either the 44-kDa or 33-kDa Pim-1 isotypes resulted in increased resistance to FLT3 inhibition-mediated cytotoxicity and apoptosis. In contrast, expression of a dominant-negative Pim-1 construct accelerated cytotoxicity in response to FLT3 inhibition and inhibited colony growth of FLT3/ITD-transformed BaF3 cells. These findings demonstrate that constitutively activated FLT3 signaling up-regulates Pim-1 expression in leukemia cells. This up-regulation contributes to the proliferative and antiapoptotic pathways induced by FLT3 signaling. (Blood. 2005;105: 1759-1767)

Sign in / Sign up

Export Citation Format

Share Document