Psp140: an immunodominant antigen in the supernatant of Streptococcus pneumoniae culture

Background and Objectives: Streptococcus pneumoniae causes many lethal infections. Due to its reduced sensitivity to commonly used antibiotics, development of new strategies against pneumococcal infections seems to be necessary. We aimed to investigate immunodominant antigens in S. pneumoniae culture supernatant in order to develop novel targets for pneumococcal vaccines. Materials and Methods: In this study S. pneumoniae ATCC49619 was sub-cultured into BHI broth from overnight culture at 37°C for 4 h. The supernatant proteins were precipitated using acetone precipitation method. A rabbit was intramuscularly immunized with alum adjuvant and 100 μg pneumococcal supernatant proteins, 6 times at 14 days' intervals to produce hyperimmune serum. ELISA assay was performed to determine the antibody level response to pneumococcal secretory proteins. Then dot blot applied for rapid evaluation of hyperimmune serum reactivity to pneumococcus supernatant proteins. The western blot was also used to determine the interaction of supernatant proteins with immunogenic rabbit's hyperimmune-serum. Results: According to the western blot analysis, the immunodominant protein had 140KDa molecular weight and designated as pneumococcal secretory protein140 (Psp140). Conclusion: The Psp140 protein in the supernatant of S. pneumoniae culture is an immunodominant protein and it is likely related to pneumococcal secretory protein or surface exposed protein which released into culture supernatant during bacterial growth.

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Use of monoclonal and polyclonal antibodies to detect prostatic acid phosphatase by immunoblotting.

Clinical Chemistry ◽

10.1093/clinchem/32.10.1832 ◽

1986 ◽

Vol 32 (10) ◽

pp. 1832-1835 ◽

Cited By ~ 2

Author(s):

P C Patel ◽

L Aubin ◽

J Côte

Keyword(s):

Monoclonal Antibodies ◽

Acid Phosphatase ◽

Western Blot ◽

Polyclonal Antibodies ◽

Prostatic Acid Phosphatase ◽

The Other ◽

Dot Blot ◽

Western Blots ◽

Dot Blot Assay ◽

Polyclonal Antisera

Abstract We investigated two techniques of immunoblotting--the Western blot and the dot blot--for use in detecting prostatic acid phosphatase (PAP, EC 3.1.3.2). We used polyclonal antisera to human PAP, produced in rabbits by hyperimmunization with purified PAP, and PAP-specific monoclonal antibodies in the immunoenzymatic protocols. We conclude that PAP can be readily detected by Western blots with use of polyclonal antisera, but not with monoclonal antibodies. On the other hand, using a dot blot assay, we could easily detect PAP with both polyclonal and monoclonal antibodies.

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Antimicrobial resistance profile and multidrug resistance patterns of Streptococcus pneumoniae isolates from patients suspected of pneumococcal infections in Ethiopia

Annals of Clinical Microbiology and Antimicrobials ◽

10.1186/s12941-021-00432-z ◽

2021 ◽

Vol 20 (1) ◽

Author(s):

Bekele Sharew ◽

Feleke Moges ◽

Gizachew Yismaw ◽

Wondwossen Abebe ◽

Surafal Fentaw ◽

...

Keyword(s):

Multidrug Resistance ◽

Streptococcus Pneumoniae ◽

Antimicrobial Resistance ◽

Addis Ababa ◽

Pneumococcal Infection ◽

Test Method ◽

Resistance Testing ◽

Global Public Health ◽

Pneumococcal Infections ◽

Resistance Patterns

Abstract Background Antimicrobial-resistant strains of Streptococcus pneumoniae have become one of the greatest challenges to global public health today and inappropriate use of antibiotics and high level of antibiotic use is probably the main factor driving the emergence of resistance worldwide. The aim of this study is, therefore, to assess the antimicrobial resistance profiles and multidrug resistance patterns of S. pneumoniae isolates from patients suspected of pneumococcal infections in Ethiopia. Methods A hospital-based prospective study was conducted from January 2018 to December 2019 at Addis Ababa city and Amhara National Region State Referral Hospitals. Antimicrobial resistance tests were performed from isolates of S. pneumoniae that were collected from pediatric and adult patients. Samples (cerebrospinal fluid, blood, sputum, eye discharge, ear discharge, and pleural and peritoneal fluids) from all collection sites were initially cultured on 5% sheep blood agar plates and incubated overnight at 37 °C in a 5% CO2 atmosphere. Streptococcus pneumoniae was identified and confirmed by typical colony morphology, alpha-hemolysis, Gram staining, optochin susceptibility, and bile solubility test. Drug resistance testing was performed using the E-test method according to recommendations of the Clinical and Laboratory Standards Institute. Results Of the 57 isolates, 17.5% were fully resistant to penicillin. The corresponding value for both cefotaxime and ceftriaxone was 1.8%. Resistance rates to erythromycin, clindamycin, tetracycline, chloramphenicol and trimethoprim-sulfamethoxazole were 59.6%, 17.5%, 38.6%, 17.5 and 24.6%, respectively. Multidrug resistance (MDR) was seen in 33.3% isolates. The most common pattern was co-resistance to penicillin, erythromycin, clindamycin, and tetracycline. Conclusions Most S. pneumoniae isolates were susceptible to ceftriaxone and cefotaxime. Penicillin has been used as a drug of choice for treating S. pneumoniae infection. However, antimicrobial resistance including multidrug resistance was observed to several commonly used antibiotics including penicillin. Hence, it is important to periodically monitor the antimicrobial resistance patterns to select empirical treatments for better management of pneumococcal infection.

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Effects of Amoxicillin Subinhibitory Concentrations on the Cross-Protection Developed by Pneumococcal Antibodies in Mouse Sepsis Caused by an Amoxicillin-Resistant Serotype 6B Streptococcus pneumoniae Strain

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.48.11.4144-4147.2004 ◽

2004 ◽

Vol 48 (11) ◽

pp. 4144-4147 ◽

Cited By ~ 3

Author(s):

D. Tarragó ◽

L. Aguilar ◽

M. J. Giménez ◽

A. Fenoll ◽

J. Casal

Keyword(s):

Streptococcus Pneumoniae ◽

Serum Levels ◽

Cross Protection ◽

Significant Activity ◽

Hyperimmune Serum ◽

Serum Peak ◽

Null Value ◽

Subinhibitory Concentrations ◽

Antibiotic Efficacy

ABSTRACT A model of mouse sepsis caused by a serotype 6B Streptococcus pneumoniae strain (amoxicillin MIC of 8 μg/ml) was developed to investigate the therapeutic effect of an amoxicillin dose (3.12 mg/kg of body weight three times daily for 48 h) producing, over the whole treatment period, subinhibitory concentrations in serum (peak concentration [C max]: 6.1 μg/ml) in animals that prior to infection had been passively immunized with a 6B or 23F hyperimmune serum (obtained by immunization with a whole-cell heat-inactivated inoculum and diluted to produce no protective effect by itself). Mortality in nonimmunized animals treated with antibiotic (3.12 mg/kg) was 90%, and mortality in animals immunized but not treated with the antibiotic was 100%. Antibiotic treatment in immunized animals produced mortality rates ≤20% when the hyperimmune serum was used, thus showing cross-protection and synergism (defined as the situation in which there is no response to the single agents [no differences versus placebo] while the combination exhibits significant activity) with subinhibitory concentrations of the antibiotic. The presence of antipneumococcal antibodies allowed antibiotic efficacy with negligible values of pharmacodynamic parameters (C max/MIC ratio of <1 and thus a null value for the time that serum levels exceed the MIC). This in vivo synergism offers a potential therapeutic strategy against resistant strains.

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Prolonged Pneumococcal Meningitis Due to an Organism With Increased Resistance to Penicillin

PEDIATRICS ◽

10.1542/peds.58.3.378 ◽

1976 ◽

Vol 58 (3) ◽

pp. 378-381 ◽

Cited By ~ 1

Author(s):

Abel Paredes ◽

Larry H. Taber ◽

Martha D. Yow ◽

Dorothy Clark ◽

William Nathan

Keyword(s):

Case Report ◽

Streptococcus Pneumoniae ◽

Pneumococcal Meningitis ◽

Pneumococcal Infections ◽

In Vitro Susceptibility ◽

Focus Of Infection ◽

Resistance To Penicillin ◽

Penicillin Treatment ◽

Susceptibility Tests

For more than 30 years, penicillin has been the agent of choice for pneumococcal infections. During this time the majority of strains of Streptococcus pneumoniae have been highly susceptible to penicillin. However, during the last ten years there have been sporadic reports of pneumococci with increased resistance to penicillin. The case report of an 18-month-old white boy with meningitis due to a strain of S. pneumoniae with increased resistance to penicillin is presented. The MIC of the organism to penicillin was 0.2µg/ml and the MBC 0.39µg/ml. The patient had normal immunity and no demonstrable sequestered focus of infection but failed to respond to appropriate doses of intravenous penicillin. Treatment with chloramphenicol caused a dramatic bacteriologic and clinical response. This experience reemphasizes the existence of pneumococcal strains of intermediate penicillin sensitivity and the importance of in vitro susceptibility tests.

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Dynamic tracking and identification of tissue-specific secretory proteins in the circulation of live mice

10.21203/rs.3.rs-90987/v1 ◽

2020 ◽

Author(s):

Jae Myoung Suh ◽

Kwang-eun Kim ◽

Isaac Park ◽

Jeesoo Kim ◽

Myeong-Gyun Kang ◽

...

Keyword(s):

Blood Plasma ◽

Mouse Liver ◽

Secretory Pathway ◽

Secretory Protein ◽

Protein Labeling ◽

Secretory Proteins ◽

Tissue Specific ◽

Dynamic Tracking

Abstract Here we describe iSLET (in situ Secretory protein Labeling via ER-anchored TurboID) which labels secretory pathway proteins as they transit through the ER-lumen to enable dynamic tracking of tissue-specific secreted proteomes in vivo. We expressed iSLET in the mouse liver and demonstrated efficient in situ labeling of the liver-specific secreted proteome which could be tracked and identified within circulating blood plasma. iSLET is a versatile and powerful tool for studying spatiotemporal dynamics of secretory proteins, a valuable class of biomarkers and therapeutic targets.

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SEROTYPING OF STREPTOCOCCUS PNEUMONIAE STRAINS, ISOLATED FROM CHILDREN IN URAL REGION WITH THE USE OF MULTIPLEX PCR

Epidemiology and Infectious Diseases (Russian Journal) ◽

10.17816/eid40701 ◽

2012 ◽

Vol 17 (5) ◽

pp. 26-30

Author(s):

E. V. Samatova ◽

A. E. Druy ◽

G. A. Tsaur ◽

L. G. Boronina

Keyword(s):

Streptococcus Pneumoniae ◽

Multiplex Pcr ◽

Inflammatory Diseases ◽

Ural Region ◽

Coincidence Rate ◽

Pneumococcal Infections ◽

Conjugate Vaccines

This article presents results of the multiplex PCR investigation of the serotypes distribution of S. pneumoniae strains circulating in Ekaterinburg and the Sverdlovsk region. This study was performed in children with invasive, noninvasive pneumococcal infections and carriers. 118 strains of pneumococci typed out of 129 ones (91.5%) referred to the 15 serotypes: 6A, 6B (20,8%); 23F (13,9%); 19F (11,5%); 8, 9V, 9A, 11F, 11A, 11B, 11C, 11D, 12F, 15A, 33F (11,5%); 3 (10%) 2, 15F, 17F, 22F, 23B (3,9%); 18B, 18C (3.9%), 19A (3,2%); 7F, 19B, 19C, 23A (3,2%); 5,10A (1.6%), 20 (1.6%), 14 (1, 6%); 9L, 9N, 15B, 15C (1,6%); 18F (1,6%); 18A (1.6%). Coincidence rate of serotypes S. pneumoniae, isolated from children with chronic infectious and inflammatory diseases of the lung with serotypes included into the content of the conjugate vaccines is: 7-valent - 69.3%, 10-valent - 98.2%, 11 - and 13-valent - 100%.

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Antimicrobial resistance profile and multidrug resistance patterns of Streptococcus pneumoniae isolates from patients suspected for pneumococcal infections in Ethiopia

10.21203/rs.3.rs-97782/v1 ◽

2020 ◽

Author(s):

BEKELE SHAREW ◽

Feleke Moges ◽

Gizachew Yismaw ◽

Wondiwossen Abebe ◽

Surafal Fentaw ◽

...

Keyword(s):

Multidrug Resistance ◽

Streptococcus Pneumoniae ◽

Antimicrobial Resistance ◽

Addis Ababa ◽

Pneumococcal Infection ◽

Test Method ◽

Resistance Testing ◽

Global Public Health ◽

Pneumococcal Infections ◽

Resistance Patterns

Abstract Backgrounds: Streptococcus pneumoniae is one of the leading causes of bacterial meningitis and pneumoniae in elderly people and children. Antimicrobial resistant strains of Streptococcus pneumoniae has been detected in all parts of the world and become one of the greatest challenges to global public health today. The aim of this study is therefore, to assess the antimicrobial resistance profiles and multidrug resistance patterns of S. pneumoniae isolates from patients suspected for pneumococcal infections in Ethiopia. Methods: A hospital-based prospective study was conducted from 2018 to 2019 at Addis Ababa and Amhara region referral hospitals. Antimicrobial resistance tests were performed on 57 isolates of S. pneumoniae that were collected from pediatric and adult patients. Samples (cerebrospinal fluid, blood, sputum, eye discharge, ear discharge, pleural and peritoneal fluids) from all collection sites were initially cultured onto 5 % sheep blood agar plates and incubated overnight at 370C in 5% CO2 atmosphere. S. pneumoniae was identified and confirmed by typical colony morphology, alpha-hemolysis, Gram staining, optochin susceptibility and bile solubility test. Drug resistance testing was performed using E-test method according to recommendations of the Clinical and Laboratory Standards Institute.Results: Of the 57 isolates, 17.5% were fully resistant to penicillin. Corresponding value for both cefotaxime and ceftriaxone was 1.8%. Resistance rates to erythromycin, clindamycin, tetracycline, chloramphenicol and trimethoprim-sulfamethoxazole were 59.6%, 17.5%, 38.6%, 17.5% and 24.6%, respectively. Multidrug resistance (MDR) was seen in 33.3% isolates. The most common pattern was co-resistance to penicillin, erythromycin, clindamycin and tetracycline.Conclusions: Most bacterial isolates were susceptible to Ceftriaxone and Cefotaxime. Penicillin has been used as a drug of choice for treating S. pneumoniae infection. However, antimicrobial resistance including multidrug resistance was observed to a number of commonly used antibiotics including penicillin. Hence, it is important to periodically monitor the antibiotic resistance patterns to choose empirical treatments for better management of pneumococcal infection.

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Toxin-Antitoxin Genes of the Gram-Positive Pathogen Streptococcus pneumoniae: So Few and Yet So Many

Microbiology and Molecular Biology Reviews ◽

10.1128/mmbr.00030-12 ◽

2012 ◽

Vol 76 (4) ◽

pp. 773-791 ◽

Cited By ~ 36

Author(s):

Wai Ting Chan ◽

Inma Moreno-Córdoba ◽

Chew Chieng Yeo ◽

Manuel Espinosa

Keyword(s):

Streptococcus Pneumoniae ◽

Biofilm Formation ◽

Bacterial Infections ◽

Host Cells ◽

Molecular Characteristics ◽

Type Ii ◽

Pneumococcal Infections ◽

Interesting Phenomenon ◽

Content Type ◽

Toxin Protein

SUMMARYPneumococcal infections cause up to 2 million deaths annually and raise a large economic burden and thus constitute an important threat to mankind. Because of the increase in the antibiotic resistance ofStreptococcus pneumoniaeclinical isolates, there is an urgent need to find new antimicrobial approaches to triumph over pneumococcal infections. Toxin-antitoxin (TA) systems (TAS), which are present in most living bacteria but not in eukaryotes, have been proposed as an effective strategy to combat bacterial infections. Type II TAS comprise a stable toxin and a labile antitoxin that form an innocuous TA complex under normal conditions. Under stress conditions, TA synthesis will be triggered, resulting in the degradation of the labile antitoxin and the release of the toxin protein, which would poison the host cells. The three functional chromosomal TAS fromS. pneumoniaethat have been studied as well as their molecular characteristics are discussed in detail in this review. Furthermore, a meticulous bioinformatics search has been performed for 48 pneumococcal genomes that are found in public databases, and more putative TAS, homologous to well-characterized ones, have been revealed. Strikingly, several unusual putative TAS, in terms of components and genetic organizations previously not envisaged, have been discovered and are further discussed. Previously, we reported a novel finding in which a unique pneumococcal DNA signature, the BOX element, affected the regulation of the pneumococcalyefM-yoeBTAS. This BOX element has also been found in some of the other pneumococcal TAS. In this review, we also discuss possible relationships between some of the pneumococcal TAS with pathogenicity, competence, biofilm formation, persistence, and an interesting phenomenon called bistability.

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An antibody against secretogranin I (chromogranin B) is packaged into secretory granules.

The Journal of Cell Biology ◽

10.1083/jcb.109.1.17 ◽

1989 ◽

Vol 109 (1) ◽

pp. 17-34 ◽

Cited By ~ 73

Author(s):

P Rosa ◽

U Weiss ◽

R Pepperkok ◽

W Ansorge ◽

C Niehrs ◽

...

Keyword(s):

Secretory Pathway ◽

Secretory Granules ◽

Intracellular Localization ◽

Secretory Protein ◽

Secretory Proteins ◽

Chromogranin B ◽

Double Labeling ◽

Protein Protein Interaction ◽

And Function ◽

Secretogranin I

We have investigated the sorting and packaging of secretory proteins into secretory granules by an immunological approach. An mAb against secretogranin I (chromogranin B), a secretory protein costored with various peptide hormones and neuropeptides in secretory granules of many endocrine cells and neurons, was expressed by microinjection of its mRNA into the secretogranin I-producing cell line PC12. An mAb against the G protein of vesicular stomatitis virus--i.e., against an antigen not present in PC12 cells--was expressed as a control. The intracellular localization and the secretion of the antibodies was studied by double-labeling immunofluorescence using the conventional and the confocal microscope, as well as by pulse-chase experiments. The secretogranin I antibody, like the control antibody, was transported along the secretory pathway to the Golgi complex. However, in contrast to the control antibody, which was secreted via the constitutive pathway, the secretogranin I antibody formed an immunocomplex with secretogranin I, was packaged into secretory granules, and was released by regulated exocytosis. Our results show that a constitutive secretory protein, unaltered by genetic engineering, can be diverted to the regulated pathway of secretion by its protein-protein interaction with a regulated secretory protein. The data also provide the basis for immunologically studying the role of luminally exposed protein domains in the biogenesis and function of regulated secretory vesicles.

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Hybridoma Screening by Antigen Capture: Reverse Dot Blot

Cold Spring Harbor Protocols ◽

10.1101/pdb.prot103135 ◽

2022 ◽

Vol 2022 (1) ◽

pp. pdb.prot103135

Author(s):

Edward A. Greenfield

Keyword(s):

Tissue Culture ◽

Monoclonal Antibodies ◽

Culture Supernatant ◽

Membrane Surface ◽

Nitrocellulose Membrane ◽

Dot Blot ◽

Tissue Culture Supernatant ◽

Antigen Capture ◽

Mouse Immunoglobulin ◽

Reverse Dot Blot

A dot blot is widely used to determine the productivity of a given hybridoma. This assay can also be used to screen a fusion or subclone plate for productive hybridoma clones. First, a nitrocellulose membrane is coated with an affinity-purified goat or rabbit anti-mouse immunoglobulin and then incubated with hybridoma tissue culture supernatant. Monoclonal antibodies in the supernatant are then “captured” on the coated nitrocellulose membrane surface and detected by screening with horseradish peroxidase (HRP).

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