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PubMed2000

Expression and purification of histidine-tagged proteins from the gram-positive Streptococcus gordonii SPEX system.

Streptococcus gordonii (S. gordonii) has been used as a gram-positive bacterial expression vector for secreted or surface-anchored recombinant proteins. Fusion of the gram-positive bacterial N-terminal signal sequence to the target protein is all that is required for efficient export. This system is termed SPEX for Surface Protein EXpression and has been used to express proteins for a variety of uses. In this study, the SPEX system has been further developed by the construction of vectors that express polyhistidine-tagged fusion proteins. SPEX vectors were constructed with an N-terminal or C-terminal histidine tag. The C-repeat region (CRR) from Streptococcus pyogenes M6 protein and the Staphylococcus aureus nuclease A (NucA) enzyme were tested for expression. The fusion proteins were purified using metal affinity chromatography (MAC). Results show that the fusion proteins were expressed and secreted from S. gordonii with the His tag at either the N- or C-terminal position and could be purified using MAC. The M6 fusions retained immunoreactivity after expression and purification as determined by immunoblots and ELISA analyses. In addition, NucA fusions retained functional activity after MAC purification. The M6-His and NucA-His fusions were purified approximately 15- and 10-fold respectively with approximately 30% recovery of protein using MAC. This study shows that the polyhistidine tag in either the N- or C-terminal position is a viable way to purify secreted heterologous proteins from the supernatant of recombinant S. gordonii cultures. This study further illustrates the value of the SPEX system for secreted expression and purification of proteins.

Base Sequence↗
PubMed1998

Evolution of Streptococcus thermophilus bacteriophage genomes by modular exchanges followed by point mutations and small deletions and insertions.

Comparative sequence analysis of 40% of the genomes from two prototype Streptococcus thermophilus bacteriophages (lytic group I phage phi Sfi19 and the cos site containing temperate phage phi Sfi21) suggested two processes in the evolution of their genomes. In a first evolutionarily distant phase the basic genome structure was apparently constituted by modular exchanges. Over the 17-kb-long DNA segment analyzed in the present report, we observed clusters of genes with similarity to genes from Leuconostoc oenos phage L10, Lactococcus lactis phage BK5-T, and Streptococcus pneumoniae phage Dp-1. A chimeric protein was predicted for orf 1291 which showed similarity to both phage BK5-T and phage Dp-1 proteins. The very large orf 1626 gene product showed similarity to two adjacent genes from the Lactobacillus delbrueckii phage LL-H and further phage proteins (Lactococcus lactis, Bacillus subtills). The similarities were localized to distinct parts of this apparently multifunctional protein. The putative phi Sfi19 lysin showed similarity to both lysins of phages and cellular enzymes. In a second, evolutionarily more recent, phase the S, thermophilus phage genomes apparently diversified by point mutations and small deletions/insertions. Over the investigated 17-kb DNA region phi Sfi19 differed from phi Sfi21 by 10% base pair changes, the majority of which were point mutations (mainly at the third codon position), while a third of the base pair differences were contributed by small deletions/insertions. The base pair changes were unevenly distributed. Over the Leuconostoc phage-related DNA the change rate was high, while over the Lactococcus and S. pneumoniae phage-related DNA the change rate was low. We speculate that the degree of base pair change could provide relative time scales for the modular exchange reactions observed in S. thermophilus phages.

Amino Acid Sequence↗
PubMed1989

Growth and adsorption of Streptococcus mutans 6715-13 to hydroxyapatite in the presence of lactoferrin.

The growth of Streptococcus mutans 6715-13 in a rich medium (Todd Hewitt broth) was drastically reduced by addition of apo-lactoferrin (apo-Lf); this effect was bacteriostatic and reversible by saturation of Lf with iron. The influence of Lf, salivary proteins (SP) and bovine serum albumin (BSA) on the attachment of Streptococcus mutans to hydroxyapatite (HA) was successively investigated. Sorption of Lf, SP, and BSA to HA was dependent on the protein concentration and reached the end-point at about 80 mg of proteins per gram of HA. Similarly, the number of streptococci adsorbed to HA was correlated to the amount of cells available up to at least 10(7) cells per mg of HA. The adsorption of Lf, SP and BSA on HA reduced the number of attaching S. mutans cells. In particular, SP reduced the adsorption of S. mutans by 30%, whereas pre-coating of HA with apo- or iron-saturated Lf resulted in a three orders of magnitude reduction of S. mutans adsorption to HA, as demonstrated by means of different experimental procedures. The powerful adherence-inhibiting effect of apo-Lf together with its noticeable antibacterial activity towards S. mutans points to a biological significance of these phenomena also in vivo.

Adsorption↗
PubMed1991

The kinetics of inflammation and phagocytosis during bovine mastitis induced by Streptococcus agalactiae bearing the protein X.

The protein X of Streptococcus agalactiae is a surface antigen borne by a high proportion of strains isolated from bovine mastitis. We have tested the capacity of two strains of X-bearing Streptococcus agalactiae to induce mastitis in dairy cows. The reference X-strain (411.07) produced an intramammary infection with local clinical signs in the three inoculated quarters. Another X-bearing strain (443.31) of bovine origin produced infection in all 11 quarters inoculated with only 25 or 85 colony-forming units. In naive cows, strain 433.31 induced less exudation of plasma into the milk, shedding of bacteria, macroscopic alteration, and a lower somatic cell count (SCC) than did the reference strain. Only one quarter spontaneously eliminated the infection before antibiotic treatment 9 days after inoculation. The serum of all the cows contained naturally acquired or induced antibodies to the challenge strain (443.31) and possessed opsonic activity. Before inflammation occurred, the milk was almost devoid of antibody or opsonic activities. The early phase of infection was characterized by rapid multiplication of streptococci in the milk, followed by a sharp drop in bacterial counts concomitant with the onset of inflammation. Three cows immunized with protein X displayed higher SCC and bactericidal activity in milk from the inoculated quarter at the onset of inflammation than non-immunized cows. Two of the three immunized cows underwent an early and transient febrile episode and eliminated the infection.

Animals↗
PubMed1978

Hydrolysis and synthesis of ATP by membrane-bound ATPase from a motile Streptococcus.

ATPase was detected in the membranes of a motile Streptococcus. Maximal enzymic activity was observed at pH 8 and ATP/Mg2+ ratio of 2. Mn2+ and Ca2+ could replace Mg2+ to some extent. Besides ATP, GTP and ITP were substrates. The enzyme was inhibited by N,N'-dicyclohexylcarbodiimide but not by sodium azide, uncouplers or bathophenanthroline. An electrochemical gradient of protons, which was artificially imposed across the membranes of Streptococcus cells by manipulation of either the K+ diffusion potential or the transmembrane pH gradient, led to ATP synthesis. ATP synthesis was abolished by proton conductors, an inhibitor of the ATPase or an increase in the extracellular K+ concentration. A comparison between the phosphate potential and the electrochemical proton gradient showed that the data found are in agreement with a stoichiometry of 2 protons translocated per molecule ATP synthesized.

Adenosine Triphosphatases↗
PubMed1987

Characterization of polymorphonuclear leukocyte aggregation in vitro induced by heat-inactivated group B streptococcus.

We studied the aggregatory characteristics of human polymorphonuclear leukocytes (PMNs) in response to heat-inactivated group B streptococcus. PMNs suspended in physiologic salt solution do not aggregate to heat-inactivated group B streptococcus (GBS) unless the GBS is previously opsonized in autologous plasma. The aggregating activity of both opsonized GBS and activated plasma are reduced if the plasma is decomplemented before incubation with GBS. Pretreatment of PMNs with pronase inhibited opsonized GBS-induced aggregation, suggesting aggregation via cell membrane receptors for opsonic fragments of C3. Pronase pretreatment had no significant effect on aggregation induced by activated plasma or arachidonic acid. Unlike PMNs in physiologic salt solution, PMNs suspended in plasma aggregate when stimulated by unopsonized GBS. GBS aggregates PMNs via complement cascade activation, opsonization, and interaction with cell membrane receptors to stimulate cellular mechanisms resulting in PMN aggregation.

Bacterial Vaccines↗
PubMed1997

Serotyping and susceptibility to macrolides and other antimicrobial drugs of Streptococcus pyogenes isolated from patients with invasive diseases in southern Israel.

Fifty-seven strains of Streptococcus pyogenes isolated from septic patients and 52 isolates from nonbacteremic patients in southern Israel were investigated for their susceptibility to new macrolides and other antimicrobial drugs. In addition, typing of the isolates by M protein and T antigen was performed. All organisms were susceptible to penicillin and chloramphenicol, 59% to tetracycline, and 7% to trimethoprim-sulfamethoxazole. All isolates but one (99%) were susceptible to clarithromycin, azithromycin, erythromycin, and clindamycin. The MIC90 of clarithromycin, erythromycin, and clindamycin was 0.064, 0.125, and 0.094 microgram/ml, respectively. Overall, 96% of the isolates could be typed by T antigen, but only 43% were M-protein typeable. No predominance of any particular M-protein type was observed. No significant differences between blood isolates and organisms derived from other sources were observed in the antibiotic susceptibility patterns or the distribution of serotypes. It is concluded that invasive Streptococcus pyogenes infections in southern Israel are caused by multiple unrelated strains. The organism remains susceptible to macrolides and clindamycin.

Anti-Bacterial Agents↗
PubMed1996

Early postsplenectomy arthritis caused by penicillin-resistant Streptococcus pneumoniae.

The first case of early-onset postsplenectomy arthritis due to intermediately penicillin-resistant Streptococcus pneumoniae is reported. After initial parenteral vancomycin and rifampin followed by oral prystinamycin-rifampin, the patient recovered without any sequelae. This case should increase awareness of the risk of invasive disease caused by penicillin-resistant Streptococcus pneumoniae in the early postoperative period after splenectomy.

Aged↗
PubMedDate not supplied

Meningitis caused by Streptococcus suis: case report and review of the literature.

A case of purulent meningitis caused by Streptococcus suis type 2 (group R streptococcus) is described. It occurred in a 69-year-old farmer's wife who raised pigs on her farm. Here, as well as in nearly all other cases of S. suis meningitis reported to date, close occupational contact with pigs or pork preceded the infection; this epidemiological link can be explained by the frequent occurrence of S. suis as a commensal and opportunistic pathogen in pigs. Up until now, S. suis infection in man has been rare and has had a good prognosis. However, disturbances of the eighth cranial nerve have been found in many patients, even causing permanent deafness in some. These and other clinical, epidemiological and microbiological features of S. suis disease in man are discussed here.

Aged↗
PubMedDate not supplied

Streptococcus agalactiae (group B) endocarditis--a description of twelve cases and review of the literature.

The group B streptococcus has been shown to be a major cause of meningitis in the newborn and an occasional cause of endocarditis and sepsis in postpartum women. Little attention has been devoted to this organism as a cause of bacterial endocarditis. Twelve patients with group B streptococcal endocarditis were seen at The Presbyterian Hospital, New York, NY, between 1974 and 1985. There were seven women, five men. Ages ranged from 32 to 81 years. Serious underlying disease was present in all - diabetes mellitus in seven, carcinoma in three (bladder in two, and breast in one), alcoholism in three, malnutrition in two, heroin addiction in one, tuberculosis in one, serious prior valvular heart disease in two. The aortic valve was affected in four patients - mitral in two, mitral and aortic in one, tricuspid in four, unknown in one. The presentation was acute in seven patients. Metastatic infection occurred in seven, heart failure in six, major emboli in four, septic pericarditis in one, myocardial abscess in one. The group B streptococcus should be considered as a pathogen capable of causing acute endocarditis in certain patients with defects of host defense, particularly patients with diabetes mellitus, carcinoma or alcoholism. Cardiac surgery may be necessary in these patients due to the rapid destruction of the valves which occurs, in spite of the fact that the organisms are usually highly susceptible to penicillin.

Adult↗
PubMed1996

Inoculum effect and bactericidal activity of cefditoren and other antibiotics against Streptococcus pneumoniae, Haemophilus influenzae, and Neisseria meningitidis.

The inoculum effect on minimum inhibitory and minimum bactericidal concentrations of cefditoren, benzylpenicillin, ampicillin, cefotaxime, ceftriaxone, and meropenem against six clinical isolates each of Streptococcus pneumoniae, Haemophilus influenzae, and Neisseria meningitidis was studied using inocula of approximately 10(4) to 10(5) and 10(7) to 10(8) cfu/ml. Vancomycin was also studied against Streptococcus pneumoniae. The inoculum effect was observed only with benzylpenicillin and ampicillin against five of six strains of Haemophilus influenzae. All antibiotics tested were bactericidal.

Anti-Bacterial Agents↗
PubMed1998

Septicemia and meningitis due to Streptococcus zooepidemicus.

A case of septicemia and meningitis due to Streptococcus zooepidemicus in an immunocompetent patient is reported. This organism is an uncommon human pathogen that sometimes causes severe infection, usually in immunocompromised patients. In the reported case, the patient required to be mechanically ventilated for one week and was treated with intravenous ampicillin and gentamicin. He recovered and was discharged from hospital three weeks after the initial presentation. Streptococcus zooepidemicus sensitive to all penicillins, was isolated from all blood cultures and the cerebrospinal fluid.

Aged↗
PubMed1994

Prevalence of antimicrobial resistance of Streptococcus pneumoniae in southwest Germany as determined by the E test.

We have studied the prevalence of anti-microbial resistance of Streptococcus pneumoniae in Southwest Germany. One hundred seventy-four clinical isolates of pneumococci collected from hospitalized patients between October 1992 and April 1994 were used for MIC determinations. MICs for penicillin, ceftriaxone, erythromycin, and rifampicin were assessed by the E test. Eleven of the 174 strains (6.3%) were intermediately resistant to penicillin (MIC between 0.1 and 1.0 microgram/ml) and four of the 174 strains (2.3%) were intermediately resistant to ceftriaxone (MIC between 0.1 and 1.0 microgram/ml). All four isolates with a reduced susceptibility to ceftriaxone also demonstrated intermediate resistance to penicillin. Six of the 174 strains (3.5%) were highly resistant (MIC > or = 8 micrograms/ml) to erythromycin. Resistance to rifampicin was not observed. Our results demonstrate that pneumococcal resistance to penicillin and erythromycin has increased markedly in Germany over the last decade. Our findings underline the need for continuous surveillance of antimicrobial resistance of Streptococcus pneumoniae.

Adult↗
PubMedDate not supplied

Bacteraemia due to different groups of beta-haemolytic streptococci: a two-year survey and presentation of a case of recurring infection due to Streptococcus "equisimilis".

We present a 2-year survey of bacteraemic episodes due to beta-haemolytic streptococci and a case of recurring infection due to group C streptococcus, Streptococcus "equisimilis". We found 53 episodes of bacteraemia with beta-haemolytic streptococci. Group A was the most common, followed by groups B, G and C. The proportion of nosocomial cases was the same in all four groups i.e. 24% (neonatal cases excluded). The clinical picture presented by groups C and G streptococcal cases was indistinguishable from that caused by group A streptococci, but patients with group G bacteraemia were older than patients with group A bacteraemia. Obvious clinical foci were more common in group A than group G cases. Disregarding neonatal cases, most patients had predisposing conditions. There was no difference in foci of bacteraemia, predisposing factors, treatment and outcome of disease. The overall mortality was 25%.

Adolescent↗
PubMed1992

Spectrum of disease in bacteraemic patients during a Streptococcus pyogenes serotype M-1 epidemic in Norway in 1988.

All 87 known cases of bacteraemia due to Streptococcus pyogenes (beta-haemolytic group A streptococci) occurring during the peak of a nationwide outbreak in Norway (population 4.2 million) between January and June 1988 were reviewed. Clinical features varied widely and appeared largely to be dependent on the patients' age. The case fatality rate ranged from 11% in the age group under 30 years to 44% in patients over 60 years. Clinical complications such as shock, severe renal or respiratory failure or serious local infection occurred particularly in 30-to 59-year old individuals. Shock was manifest in 32% of the patients and carried a 68% case fatality rate. Chronic heart disease in the elderly and pneumonia seemed to be associated with a fatal outcome. In the 25 patients (29%) who died the disease showed a fulminant course, 80% dying within 48 hours after admission. However, 56% of the patients had experienced symptoms for more than two days before admission, suggesting that early diagnosis and treatment might possibly have prevented the development of a serious disease. This study revealed a wide spectrum of clinical manifestations in bacteraemia cases in a unique epidemiological situation caused largely by a single serotype of Streptococcus pyogenes; 89% of the 27 preserved bacteraemia strains carried the M-1 antigen. The observations call attention to the ability of these organisms to cause fulminant clinical illness, indicating a probable increase in both invasiveness and toxicity of group A streptococci responsible for the epidemic.

Adolescent↗
PubMed1988

Vaginitis caused by nutritionally variant Streptococcus pyogenes.

A nutritionally variant Streptococcus pyogenes was isolated from a vaginal specimen. The organism was isolated in essentially pure culture with a few colonies of normal vaginal flora. The bacterium was identified as Streptococcus pyogenes with the use of rapid test kits and the presence of group A antigen.

Female↗
PubMed1989

Fatal Streptococcus suis septicemia in an abattoir worker.

A 51-year-old abattoir worker became ill with high fever and was admitted to hospital late in the evening, where he died 4 h later. Smears of peripheral blood taken 7 h after death showed numerous cocci. Cultures of the blood grew beta-hemolytic streptococci exhibiting the biochemical properties of Streptococcus suis II. A complete autopsy with extensive microscopical studies showed numerous bacteria in small vessels of several organs, but no bacterial colonisation of tissues, meningitis, or other relevant lesions were found. The group R antigen of the bacteria could be demonstrated by immunofluorescence in paraffin sections of the kidney and the adrenal gland. Two similar cases of fulminant fatal Streptococcus suis II septicemia have been reported from Denmark. This appears to be the first case observed in the Federal Republic of Germany.

Abattoirs↗
PubMed1990

Identification of atypical strains of Streptococcus pneumoniae by a specific DNA probe.

A specific DNA probe containing a 0.65 Kb fragment coding for the aminoterminal region of the major pneumococcal autolysin (amidase) was constructed, deleting the region involved in the specific recognition of cell wall choline residues. The high specificity of this probe was demonstrated in tests with Streptococcus pneumoniae related species including Streptococcus oralis, which contains choline in its cell wall. The probe was used to characterize pneumococcal isolates showing atypical responses in conventional identification tests. The hybridization obtained with 27 atypical pneumococci and 11 of 17 isolates presumptively identified as viridans streptococci confirmed that the probe is suitable for diagnostic use.

DNA Probes↗