scholarly journals Carbon source regulates polysaccharide capsule biosynthesis in Streptococcus pneumoniae

2019 ◽  
Vol 294 (46) ◽  
pp. 17224-17238 ◽  
Author(s):  
Lukas J. Troxler ◽  
Joel P. Werren ◽  
Thierry O. Schaffner ◽  
Nadezda Mostacci ◽  
Peter Vermathen ◽  
...  
Keyword(s):  
Streptococcus Pneumoniae ◽  
Carbon Source ◽  
Capsular Polysaccharide ◽  
Carbon Sources ◽  
Cell Extract ◽  
Therapeutic Approaches ◽  
Capsule Production ◽  
Important Virulence Factor ◽  
Capsule Thickness ◽  
Whole Cell Extract

The exopolysaccharide capsule of Streptococcus pneumoniae is an important virulence factor, but the mechanisms that regulate capsule thickness are not fully understood. Here, we investigated the effects of various exogenously supplied carbohydrates on capsule production and gene expression in several pneumococcal serotypes. Microscopy analyses indicated a near absence of the capsular polysaccharide (CPS) when S. pneumoniae was grown on fructose. Moreover, serotype 7F pneumococci produced much less CPS than strains of other serotypes (6B, 6C, 9V, 15, and 23F) when grown on glucose or sucrose. RNA-sequencing revealed carbon source-dependent regulation of distinct genes of WT strains and capsule-switch mutants of serotypes 6B and 7F, but could not explain the mechanism of capsule thickness regulation. In contrast, 31P NMR of whole-cell extract from capsule-knockout strains (Δcps) clearly revealed the accumulation or absence of capsule precursor metabolites when cells were grown on glucose or fructose, respectively. This finding suggests that fructose uptake mainly results in intracellular fructose 1-phosphate, which is not converted to CPS precursors. In addition, serotype 7F strains accumulated more precursors than did 6B strains, indicating less efficient conversion of precursor metabolites into the CPS in 7F, in line with its thinner capsule. Finally, isotopologue sucrose labeling and NMR analyses revealed that the uptake of the labeled fructose subunit into the capsule is <10% that of glucose. Our findings on the effects of carbon sources on CPS production in different S. pneumoniae serotypes may contribute to a better understanding of pneumococcal diseases and could inform future therapeutic approaches.

scholarly journals Identification and Characterization of thecps Locus of Streptococcus suis Serotype 2: the Capsule Protects against Phagocytosis and Is an Important Virulence Factor

1999 ◽  
Vol 67 (4) ◽  
pp. 1750-1756 ◽  
Author(s):  
Hilde E. Smith ◽  
Marloes Damman ◽  
Joeke van der Velde ◽  
Frans Wagenaar ◽  
Henk J. Wisselink ◽  
...  
Keyword(s):  
Insertional Mutagenesis ◽  
Capsular Polysaccharide ◽  
Streptococcus Suis ◽  
Open Reading Frames ◽  
Transcriptional Unit ◽  
Polysaccharide Biosynthesis ◽  
Capsule Production ◽  
Important Virulence Factor ◽  
Isogenic Mutants

ABSTRACT To study the role of the capsule of Streptococcus suisserotype 2 in virulence, we generated two isogenic mutants disturbed in capsule production. For that purpose, we first cloned and characterized a major part of the capsular polysaccharide biosynthesis (cps) locus of S. suis serotype 2. Based on the established sequence, 14 open reading frames (ORFs), designated Orf2Z, Orf2Y, Orf2X, and Cps2A to Cps2K, were identified. Twelve ORFs belonged to a single transcriptional unit. The gene products of 11 of these ORFs showed similarity to proteins involved in polysaccharide biosynthesis of other gram-positive microorganisms. Nonencapsulated isogenic mutants were generated in the cps2B and cps2EF genes by insertional mutagenesis. In contrast to the wild-type S. suis serotype 2 strain, the nonencapsulated strains were highly sensitive to ingestion by porcine alveolar lung macrophages in vitro. More importantly, the nonencapsulated mutant strains were completely avirulent in young germfree pigs after intranasal inoculation. These observations indicate that the capsule of S. suis serotype 2 plays an essential role in the pathogenesis of S. suisserotype 2 infections.

scholarly journals Carbon Source-Dependent Changes of the Structure of Streptococcus pneumoniae Capsular Polysaccharide with Serotype 6F

2021 ◽  
Vol 22 (9) ◽  
pp. 4580
Author(s):  
Joel P. Werren ◽  
Lukas J. Troxler ◽  
Oluwaseun Rume-Abiola Oyewole ◽  
Alban Ramette ◽  
Silvio D. Brugger ◽  
...  
Keyword(s):  
Streptococcus Pneumoniae ◽  
Carbon Source ◽  
1H Nmr ◽  
31P Nmr ◽  
Capsular Polysaccharide ◽  
Defined Medium ◽  
Flow Cytometric ◽  
Unequivocal Identification ◽  
Pneumococcal Serotype ◽  
Polysaccharide Composition

The structure of the exopolysaccharide capsule of Streptococcus pneumoniae is defined by the genetic arrangement of the capsule operon allowing the unequivocal identification of the pneumococcal serotype. Here, we investigated the environment-dependent composition of the polysaccharide structure of S. pneumoniae serotype 6F. When grown in a chemically defined medium (CDM) with glucose versus galactose, the exopolysaccharide capsule of the serotype 6F strains reveals a ratio of 1/0.6 or 1/0.3 for galactose/glucose in the capsule by 1H-NMR analyses, respectively. Increased production of the capsule precursor UDP-glucose has been identified by 31P-NMR in CDM with glucose. Flow cytometric experiments using monoclonal antibodies showed decreased labelling of Hyp6AG4 (specific for serotype 6A) antibodies when 6F is grown in glucose as compared to galactose, which mirrors the 1H-NMR results. Whole-genome sequencing analyses of serotype 6F isolates suggested that the isolates evolved during two different events from serotype 6A during the time when the 13-valent pneumococcal conjugate vaccine (PCV-13) was introduced. In conclusion, this study shows differences in the capsular structure of serotype 6F strains using glucose as compared to galactose as the carbon source. Therefore, 6F strains may show slightly different polysaccharide composition while colonizing the human nasopharynx (galactose rich) as compared to invasive locations such as the blood (glucose rich).

Immobilization-based isolation of capsule-negative mutants of Streptococcus pneumoniae

Microbiology ◽  
2005 ◽  
Vol 151 (6) ◽  
pp. 1911-1917 ◽  
Author(s):  
Daniel Llull ◽  
Patrick Veiga ◽  
Josselyne Tremblay ◽  
Saulius Kulakauskas
Keyword(s):  
Streptococcus Pneumoniae ◽  
Capsular Polysaccharide ◽  
Agar Medium ◽  
Upper Respiratory Tract ◽  
Gene Encoding ◽  
Capsule Production ◽  
Bacterial Immobilization ◽  
Reliable Procedure

The capsular polysaccharide (CPS) is the most important identified virulence factor of Streptococcus pneumoniae, a human pathogen of the upper respiratory tract. One limitation in studies of S. pneumoniae surface virulence factors is the lack of a reliable procedure for isolation of capsule-negative mutants of clinical strains. This paper presents an approach, based on the immobilization of pneumococci in semi-liquid (0·04 % agar) medium, to easily distinguish and select for non-capsulated mutants. A clinical S. pneumoniae type 37 strain was used as a model to show that CPS production results in bacterial immobilization in semi-liquid agar medium and restricts cell sedimentation. Descendants of CPS− mutants sedimented faster under these conditions and therefore could be separated from immobilized parental cells. The CPS− phenotype of the obtained mutants was confirmed by both immunoagglutination and immunostaining experiments using specific type 37 capsular antibodies. Complementation of immobilization with the cloned tts gene, encoding type 37 CPS synthase, confirmed that faster sedimentation of mutants was specifically due to loss of the capsule. DNA sequence determination of three independent mutants revealed a point mutation, a 46 nt deletion and a heptanucleotide duplication in the tts gene. Immobilization of strains producing other CPSs (type 2, 3 and 6) also resulted in the appearance of CPS− mutants, thus showing that immobilization-based isolation is not restricted to type 37 pneumococci. Bacterial growth in semi-liquid medium proved to be a useful model system to identify the genetic consequences of immobilization. The results indicate that immobilization due to CPS may impose selective pressure against capsule production and thus contribute to capsule plasticity.

scholarly journals Capsule Prolongs Survival of Streptococcus pneumoniae during Starvation

2018 ◽  
Vol 86 (3) ◽  
Author(s):  
Shigeto Hamaguchi ◽  
M. Ammar Zafar ◽  
Michael Cammer ◽  
Jeffrey N. Weiser
Keyword(s):  
Streptococcus Pneumoniae ◽  
Capsular Polysaccharide ◽  
Close Contact ◽  
Carbon Sources ◽  
Strain Differences ◽  
Content Type ◽  
Polysaccharide Capsules ◽  
Phosphate Buffered Saline ◽  
Capsule Size

ABSTRACT Person-to-person transmission of Streptococcus pneumoniae (the pneumococcus) may occur via environmental sources in close contact with carriers. Pneumococcal polysaccharide capsules, the determinant of serotype (or type), are heterogeneous in structure and amount, and these differences affect rates of transmission. In this study, we examined the contribution of capsule and its variations to the maintenance of pneumococcal viability under starvation conditions. S. pneumoniae retained its ability to colonize infant mice even after incubation for 24 h in phosphate-buffered saline at 25°C. The expression of capsule by the cps locus prolonged survival under these and other nutrient-poor conditions. Analysis of capsule-switch constructs showed that strain-to-strain differences in survival were due to capsule type rather than genetic background. The addition of glucose was sufficient to rescue the survival defect of the capsule-deficient derivative, demonstrating that in the absence of capsule, survival depends upon nutrient availability. During starvation, there was a decrease in capsule size and amount of capsular polysaccharide that was dependent on bacterial viability and the presence of the cps locus. These observations suggest that pneumococci catabolize their own capsular polysaccharide using the genes involved in its biosynthesis to maintain viability when other carbon sources are unavailable. Our findings describe a new role of the pneumococcal capsule: the prolongation of viability under nutrient-limiting conditions as would be encountered during periods when the organism is between hosts.

scholarly journals Capsular Polysaccharide Expression in CommensalStreptococcusSpecies: Genetic and Antigenic Similarities toStreptococcus pneumoniae

mBio ◽  
2016 ◽  
Vol 7 (6) ◽  
Author(s):  
Uffe B. Skov Sørensen ◽  
Kaihu Yao ◽  
Yonghong Yang ◽  
Hervé Tettelin ◽  
Mogens Kilian
Keyword(s):  
Streptococcus Pneumoniae ◽  
Virulence Factors ◽  
Capsular Polysaccharide ◽  
Capsular Polysaccharides ◽  
Structural Polymorphism ◽  
Commensal Microbiota ◽  
Capsule Production ◽  
Genetic Mechanisms ◽  
Different Origins ◽  
Host Parasite

ABSTRACTExpression of a capsular polysaccharide is considered a hallmark of most invasive species of bacteria, includingStreptococcus pneumoniae, in which the capsule is among the principal virulence factors and is the basis for successful vaccines. Consequently, it was previously assumed that capsule production distinguishesS. pneumoniaefrom closely related commensals of the mitis group streptococci. Based on antigenic and genetic analyses of 187 mitis group streptococci, including 90 recognized serotypes ofS. pneumoniae, we demonstrated capsule production by the Wzy/Wzx pathway in 74% of 66S. mitisstrains and in virtually all tested strains ofS. oralis(subspeciesoralis,dentisani, andtigurinus) andS. infantis. Additional analyses of genomes ofS. cristatus,S. parasanguinis,S. australis,S. sanguinis,S. gordonii,S. anginosus,S. intermedius, andS. constellatusrevealed complete capsular biosynthesis (cps) loci in all strains tested. Truncatedcpsloci were detected in three strains ofS. pseudopneumoniae, in 26% ofS. mitisstrains, and in a singleS. oralisstrain. The level of sequence identities ofcpslocus genes confirmed that the structural polymorphism of capsular polysaccharides inS. pneumoniaeevolved by import ofcpsfragments from commensalStreptococcusspecies, resulting in a mosaic of genes of different origins. The demonstrated antigenic identity of at least eight of the numerous capsular polysaccharide structures expressed by commensal streptococci with recognized serotypes ofS. pneumoniaeraises concerns about potential misidentifications in addition to important questions concerning the consequences for vaccination and host-parasite relationships both for the commensals and for the pathogen.IMPORTANCEExpression of a capsular polysaccharide is among the principal virulence factors ofStreptococcus pneumoniaeand is the basis for successful vaccines against infections caused by this important pathogen. Contrasting with previous assumptions, this study showed that expression of capsular polysaccharides by the same genetic mechanisms is a general property of closely related species of streptococci that form a significant part of our commensal microbiota. The demonstrated antigenic identity of many capsular polysaccharides expressed by commensal streptococci andS. pneumoniaeraises important questions concerning the consequences for vaccination and host-parasite relationships both for the commensals and the pathogen.

scholarly journals Positive Correlation between Tyrosine Phosphorylation of CpsD and Capsular Polysaccharide Production in Streptococcus pneumoniae

2003 ◽  
Vol 185 (20) ◽  
pp. 6057-6066 ◽  
Author(s):  
Matthew H. Bender ◽  
Robert T. Cartee ◽  
Janet Yother
Keyword(s):  
Cell Wall ◽  
Streptococcus Pneumoniae ◽  
Tyrosine Phosphorylation ◽  
Capsular Polysaccharide ◽  
Kinase Inhibitor ◽  
Tyrosine Phosphatase ◽  
Regulatory System ◽  
Capsule Production ◽  
Capsule Locus ◽  
Ladder Pattern

ABSTRACT CpsA, CpsB, CpsC, and CpsD are part of a tyrosine phosphorylation regulatory system involved in modulation of capsule synthesis in Streptococcus pneumoniae and many other gram-positive and gram-negative bacteria. Using an immunoblotting technique, we observed distinct laddering patterns of S. pneumoniae capsular polysaccharides of various serotypes and found that transfer of the polymer from the membrane to the cell wall was independent of size. Deletion of cps2A, cps2B, cps2C, or cps2D in the serotype 2 strain D39 did not affect the ability to transfer capsule to the cell wall. Deletion of cps2C or cps2D, which encode two domains of an autophosphorylating tyrosine kinase, resulted in the production of only short-chain polymers. The function of Cps2A is unknown, and the polymer laddering pattern of the cps2A deletion mutants appeared similar to that of the parent, although the total amount of capsule was decreased. Loss of Cps2B, a tyrosine phosphatase and a kinase inhibitor, resulted in an increase in capsule amount and a normal ladder pattern. However, Cps2B mutants exhibited reduced virulence following intravenous inoculation of mice and were unable to colonize the nasopharynx, suggesting a diminished capacity to sense or respond to these environments. In D39 and its isogenic mutants, the amounts of capsule and tyrosine-phosphorylated Cps2D (Cps2D∼P) correlated directly. In contrast, restoration of type 2 capsule production followed by deletion of cps2B in Rx1, a laboratory passaged D39 derivative containing multiple uncharacterized mutations, resulted in decreased capsule amounts but no alteration in Cps2D∼P levels. Thus, a factor outside the capsule locus, which is either missing or defective in the Rx1 background, is important in the control of capsule synthesis.

Effect of carbon source on Phenobarbital metabolism by Rhizopus stolonifer

2010 ◽  
Vol 3 (2) ◽  
pp. 1022-1027
Author(s):  
Kavitha K ◽  
Asha S ◽  
Hima Bindu T.V.L ◽  
Vidyavathi M
Keyword(s):  
Carbon Source ◽  
High Performance ◽  
Carbon Sources ◽  
In Vitro Models ◽  
Rhizopus Stolonifer ◽  
Safety And Efficacy ◽  
Alternative Systems ◽  
Toxicological Studies ◽  
Microbial Model

The safety and efficacy of a drug is based on its metabolism or metabolite formed. The metabolism of drugs can be studied by different in vitro models, among which microbial model became popular. In the present study, eight microbes were screened for their ability to metabolize phenobarbital in a manner comparable to humans with a model to develop alternative systems to study human drug metabolism. Among the different microbes screened, a filamentous fungi Rhizopus stolonifer metabolized phenobarbital to its metabolite which is used for further pharmacological and toxicological studies. The transformation of phenobarbital was identified by high- performance liquid chromatography (HPLC). Interestingly, Rhizopus stolonifer sample showed an extra metabolite peak at 3.11min. compared to its controls. The influence of different carbon sources in media used for growth of fungus, on metabolite production was studied, to find its effect in production of metabolite as the carbon source may influence the growth of the cell.

scholarly journals Denitrification Control in a Recirculating Aquaculture System—A Simulation Study

Processes ◽  
2020 ◽  
Vol 8 (10) ◽  
pp. 1306
Author(s):  
Pedro Almeida ◽  
Laurent Dewasme ◽  
Alain Vande Wouwer
Keyword(s):  
Carbon Source ◽  
Carbon Sources ◽  
Aquatic Organisms ◽  
Operating Conditions ◽  
Toxic Substances ◽  
Recirculating Aquaculture System ◽  
Treatment Technology ◽  
Tuning Method ◽  
Recirculating Aquaculture ◽  
Aquaculture System

The recirculating aquaculture system (RAS) is a land-based water treatment technology, which allows for farming aquatic organisms, such as fish, by reusing the water in the production (often less than 5%). This technology is based on the use of filters, either mechanical or biological, and can, in principle, be used for any species grown in aquaculture. Due to the low recirculation rate, ammonia accumulates in the system and must be converted into nitrate using nitrification reactors. Although less toxic for fish, nitrate can also be further reduced into nitrogen gas by the use of denitrification biofilters which may create several issues, such as incomplete denitrification, resulting in toxic substances, such as nitrite and nitric oxide, or a waste of carbon source in excess. Control of the added quantity of carbon source in the denitrification biofilter is then mandatory to keep nitrate/nitrite concentrations under toxic levels for fish and in accordance with local effluent regulations, and to reduce costs related to wasted organic carbon sources. This study therefore investigates the application of different control methodologies to a denitrification reactor in a RAS. To this end, a numerical simulator is built to predict the RAS behavior and to allow for the comparison of different control approaches, in the presence of changes in the operating conditions, such as fish density and biofilter removal efficiency. First, a classical proportional-integral-derivative (PID) controller was designed, based on an SIMC tuning method depending on the amount of ammonia excreted by fish. Then, linearizing and cascade controllers were considered as possible alternatives.

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