Comparison of bacterial flora in the oral cavity of children and adults

The aim of the study was to assess levels of occurrence and number of aerobic bacteria hemolysing and non-hemolysing, anaerobic bacteria, streptococci hemolysing and non-hemolysing, staphylococci, and bacteria responsible for tooth decay (Streptococcus mutans and Lactobacillus spp.) on oral cavity in children and adults. The results obtained indicate the difference of the level of occurrence of, aerobic bacteria hemolysing and non-hemolysing, anaerobic bacteria, streptococci hemolysing and non-hemolysing, staphylococci, and bacteria responsible for tooth decay (Streptococcus mutans and Lactobacillus spp.) was not statistically significant in either group. The counts and average values of the counts for aerobic bacteria non-hemolysing, anaerobic bacteria, streptococci hemolysing and non-hemolysing and Streptococcus mutans turned out to be statistically significantly larger in adults than in children. However for aerobic bacteria hemolysing, staphylococci and Lactobacillus spp. the difference of the counts was not statistically significant in either group.     

天然药物蜂房化学成分提取物对口腔细菌生长的实验研究

目的研究蜂房中分离得到的不同组分对致龋菌生长的影响,寻找蜂房抑龋的有效成分。方法通过溶剂分段和层析技术对蜂房进行分离,得到4个组分,采用液体稀释法研究蜂房不同组分对口腔常居细菌——血液链球菌、唾液链球菌,以及4种主要致龋菌——变形链球菌、内氏放线菌、粘性放线菌和乳酸杆菌生长的影响,使用活菌计数法测量五倍子总鞣质及各组分对变形链球菌生长曲线的影响。结果蜂房提取物中1、2和3组分对实验菌有较强的抑菌作用,蜂房提取物3组分对于变形链球菌生长曲线的抑制作用最强。结论蜂房各组分对实验菌都有一定的抑菌作用,其抑菌作用可能和其中的甾醇类化合物有关。     

A high incidence of prophage carriage among natural isolates of Streptococcus pneumoniae.

The majority (591 of 791, or 76%) of Streptococcus pneumoniae clinical isolates examined showed the presence of two or more chromosomal SmaI fragments that hybridized with the lytA-specific DNA probe. Only one of these fragments, frequently having an approximate molecular size of 90 kb, was shown to carry the genetic determinant of the pneumococcal autolysin (N-acetylmuramic acid-L-alanine amidase). Strains carrying multiple copies of lytA homologues included both antibiotic-susceptible and -resistant isolates as well as a number of different serotypes and strains recovered from geographic sites on three continents. Mitomycin C treatment of strains carrying several lytA-hybridizing fragments caused the appearance of extrachromosomal DNA hybridizing to the lytA gene, followed by lysis of the bacteria. Such lysates contained phage particles detectable by electron microscopy. The findings suggest that the lytA-hybridizing fragments in excess of the host lytA represent components of pneumococcal bacteriophages. The high proportion of clinical isolates carrying multiple copies of lytA indicates the widespread occurrence of lysogeny, which may contribute to genetic variation in natural populations of pneumococci.     

Autophagy vs. Group A Streptococcus

We have identified several mammalian protein components of the autophagy pathway. By using them as powerful tools to examine the functional significance of this degradation pathway, we recently showed that autophagy efficiently kills a pathogenic bacterium, Group A Streptococcus, after it invades host cells. However, the autophagosomes induced by these bacteria have features distinct from those of the canonical pathway.     

Autophagy vs. Group A Streptococcus

We have identified several mammalian protein components of the autophagy pathway. By using them as powerful tools to examine the functional significance of this degradation pathway, we recently showed that autophagy efficiently kills a pathogenic bacterium, Group A Streptococcus, after it invades host cells. However, the autophagosomes induced by these bacteria have features distinct from those of the canonical pathway.     

Microbial spoilage of pre-cooked potato-topped pies

The ecological succession of bacteria which developed in pre-cooked potato-topped pies stored at two different temperatures was examined. Bacillus, Streptococcus and Staphylococcus-Micrococcus spp. were the predominant organisms isolated from freshly prepared pies and those stored at 4°C and 37°C. None of these groups of bacteria caused significant biodeterioration of pies held at 4°C, but all groups grew well in pies stored at 37°C and achieved counts of ca 10⁸/g of sample. Bacillus spp. were the first group to grow, followed by Streptococcus and Staphylococcus- Micrococcus spp. Growth which occurred at 37°C did so at the expense of glucose, lactate accumulated and the pH of pie components decreased. Amylase activity detected in all pie components during storage was associated with the growth of Bacillus spp. and probably supplemented glucose already present in pies, by hydrolytic cleavage of potato, flour or binder starches. Spoilage caused by growth and activity of the bacteria isolated was not associated with visual signs of bio-deterioration, nor production of 'off' odour usually associated with spoilage of meats. These results suggest that pre-cooked potato-topped pies held at inappropriate temperatures represent a potential public health risk.     

Microbial spoilage of pre-cooked potato-topped pies

The ecological succession of bacteria which developed in pre-cooked potato-topped pies stored at two different temperatures was examined. Bacillus, Streptococcus and Staphylococcus-Micrococcus spp. were the predominant organisms isolated from freshly prepared pies and those stored at 4 degrees and 37 degrees C. None of these groups of bacteria caused significant biodeterioration of pies held at 4 degrees C, but all groups grew well in pies stored at 37 degrees C and achieved counts of ca 10(8)/g of sample. Bacillus spp. were the first group to grow, followed by Streptococcus and Staphylococcus-Micrococcus spp. Growth which occurred at 37 degrees C did so at the expense of glucose, lactate accumulated and the pH of pie components decreased. Amylase activity detected in all pie components during storage was associated with the growth of Bacillus spp. and probably supplemented glucose already present in pies, by hydrolytic cleavage of potato, flour or binder starches. Spoilage caused by growth and activity of the bacteria isolated was not associated with visual signs of biodeterioration, nor production of 'off' odour usually associated with spoilage of meats. These results suggest that pre-cooked potato-topped pies held at inappropriate temperatures represent a potential public health risk.     

Predominant Catalase-negative Soil Bacteria. I. Streptococcal Population Indigenous to Soil

Streptococci related to Streptococcus sanguis were shown to represent a major segment of the bacterial flora of certain soils. Upon initial isolation, these streptococci were quite pleomorphic and displayed morphological characteristics common to several other genera of bacteria, but after prolonged cultivation on laboratory media the isolates demonstrated a typical streptococcal morphology. It was proposed that a requirement for correction of the control mechanisms affecting growth rate and cell wall synthesis in part might explain the difficulty encountered in isolation of these organisms from soil and their initial pleomorphism and cultural instability. The possible role of these soil organisms as etiological agents of disease and the occurrence of other groups of streptococci in soil were discussed.     

Use of leukocytes as treatment for endometritis in mares experimentally infected with Streptococcus equi subsp. zooepidemicus

This study compared four treatments for bacterial endometritis in mares experimentally infected with Streptococcus zooepidemicus. Twenty-five mares were used, 20 resistant and five susceptible to endometritis. Mares would be in estrus when infected. Twenty-four hours after inoculation, clinical, bacteriological and cytological examinations were performed and repeated until the first occurrence: negative cytology and no Streptococcus growth or the seventh day post-infection. All mares showed clinical signs of endometritis and were assigned to one of the following treatments: (1) intrauterine infusion of fresh leukocytes; (2) intrauterine infusion of frozen-thawed leukocytes; (3) intrauterine infusion of lysed leukocytes; (4) intrauterine infusion of recombinant human interleukin-8 (rhIL-8); (5) control. Mares were submitted to all treatments, with at least a 14-day interval between treatments in a Latin square design. Treatment did not affect (P=0.121) time needed for resistant mares to eliminate bacteria. Time needed for elimination of bacteria was similar in susceptible mares treated with fresh and frozen leukocytes (P=0.333). Susceptible mares treated with frozen leukocytes also did not differ from those treated with lysed leukocytes (P=0.227) for time to eliminate bacteria, but were significantly different (P>0.02) from those treated with rhIL-8 and control. In resistant mares, physical clearance ability was probably the responsible for bacterial elimination. Intrauterine infusions in susceptible mares with viable or lysed leukocytes associated or not to opsonizing factors, reduced the time to elimination of bacteria. Infusions with bactericidal effect (functional neutrophils and granules) was likely effective and responsible for the more rapid elimination of bacteria in susceptible mares.     

Cutting edge: recognition of Gram-positive bacterial cell wall components by the innate immune system occurs via Toll-like receptor 2.

Invasive infection with Gram-positive and Gram-negative bacteria often results in septic shock and death. The basis for the earliest steps in innate immune response to Gram-positive bacterial infection is poorly understood. The LPS component of the Gram-negative bacterial cell wall appears to activate cells via CD14 and Toll-like receptor (TLR) 2 and TLR4. We hypothesized that Gram-positive bacteria might also be recognized by TLRs. Heterologous expression of human TLR2, but not TLR4, in fibroblasts conferred responsiveness to Staphylococcus aureus and Streptococcus pneumoniae as evidenced by inducible translocation of NF-kappaB. CD14 coexpression synergistically enhanced TLR2-mediated activation. To determine which components of Gram-positive cell walls activate Toll proteins, we tested a soluble preparation of peptidoglycan prepared from S. aureus. Soluble peptidoglycan substituted for whole organisms. These data suggest that the similarity of clinical response to invasive infection by Gram-positive and Gram-negative bacteria is due to bacterial recognition via similar TLRs.     

Deoxyribonucleic acid sequence common to staphylococcal and streptococcal plasmids which specify erythromycin resistance.

Plasmids from erythromycin-resistant Staphylococcus aureus, Streptococcus sanguis, and Streptococcus faecalis show deoxyribonucleic acid sequence homology. The homologous sequences can be localized to specific restriction endonuclease fragments, which in the case of S. aureus plasmid pI258 involves a single fragment from either EcoRI or HindIII digest known to contain the erythromycin resistance determinant. Complementary ribonucleic acid probes prepared from S. aureus plasmid pI258 and S. sanguis plasmid pAM77 also hybridize to specific fragments in restriction endonuclease digests of deoxyribonucleic acid from erythromycin-resistant Streptococcus progenes and Streptococcus pneumoniae. These studies suggest a common origin for a class of erythromycin resistance determinants in unrelated strains of pathogenic bacteria for which exchange of genetic material has not been demonstrated.     

Preservation of capsular material of streptococcal cells by specific lectins determined by immunoelectron microscopy

Summary We describe the use of lectins as specific stabilizing agents for the polysaccharide capsular components of two Gram-positive bacteria,Streptococcus agalactiae andStreptococcus bovis. Treatment of bacterial suspensions with wheatgerm agglutinin and concanavalin A allowed better morphological preservation as well as immunoelectron microscopic localization of a capsular component (lipoteichoic acid) by employing specific antibodies and the protein A-gold technique. Data obtained indicate that lectins are useful agents in preserving highly water-soluble capsular components during the electron microscopy procedures for both unembedded and embedded samples.     

A novel resistance mechanism against beta-lactams in Streptococcus pneumoniae involves CpoA, a putative glycosyltransferase.

Piperacillin resistance in Streptococcus pneumoniae was mediated by mutations in a novel gene, cpoA, that also confer transformation deficiency and a decrease in penicillin-binding protein la. cpoA is part of an operon located downstream of the primary sigma factor of S. pneumoniae. The deduced protein, CpoA, and the peptide encoded by the adjacent 3' open reading frame contained domains homologous to glycosyltransferases of procaryotes and eucaryotes that act on membrane-associated substrates, such as enzymes functioning in lipopolysaccharide core biosynthesis of gram-negative bacteria, RodD of Bacillus subtilis, which is involved in teichoic acid biosynthesis, and the human PIG-A protein, which is required for early steps of glycosylphosphatidylinositol anchor biosynthesis. This suggests that the cpo operon has a similar function related to cell surface components.     

Aerobic bacteria from nasal discharge of goats suffering from clinical PPR: isolation and identification

Bacteriological isolation of aerobic bacteria from nasal discharge of achondroplastic West African dwarf goats showing clinical signs of peste des petits ruminants (PPR), and the laboratory procedures carried out for the identification of the isolates are described. From a total of forty nasal swabs obtained seventy five isolates were recovered. The percentage frequency of isolation of different organisms in descending order of occurrence is as follows: Staphylococcus 44.0%; Streptococcus 22.67%; Neisseria 12.00%; Pasteurella 10.67%; Pseudomonas 4.00%; Proteus 4.00% and Corynebacterium 1.33%. A further break-down using biochemical tests revealed that Staphylococcus aureus accounted for 30.67% of the total bacteria isolates; Staphylococcus epidermidis 13.33%; Streptococcus viridans 18.67%; Streptococcus pyogenes 4.00%; Neisseria catarrhalis 12.00%; Proteus mirabilis 2.67%; Proteus rettgeri 1.33%; Pseudomonas aeruginosa 5.33%; Pasteurella multocida 4.00%; Pasteurella haemolytica 6.67% and Corynebacterium pyogenes 1.33%.     

Energy transduction in the bacterial flagellar motor. Effects of load and pH.

The effect of load and pH on the relation between proton potential and flagellar rotation has been studied in cells of a smooth-swimming Streptococcus strain. The driving potential, speeds of free-swimming bacteria, and rotation rates of bacteria tethered to glass by a single flagellum were measured. The relation between rotation rate of tethered bacteria and potential was remarkably linear up to nearly -200 mV. The relation between swimming speed and potential exhibited both saturation and threshold, as previously observed in other species. The form of these relations depended on pH. The equivalence of the electrical and chemical potential components of the proton potential in enabling swimming depended on the voltage. Our observations may be most simply accommodated by a kinetic scheme that links transmembrane proton transits to a tightly coupled work cycle. The properties of this scheme were elucidated by computer simulations of the experimental plots. These simulations indicated that the protonable groups that participate in the rate limiting reactions have a fractional electrical distance between three-fourths to all of the way toward the cytoplasm with a corresponding mean proton binding affinity of 10(-7.3)-10(-7.0) M, respectively.     

Interbacterial adhesion between Pseudomonas aeruginosa and indigenous oral bacteria isolated from patients with cystic fibrosis

Interbacterial adhesion between strains of Pseudomonas aeruginosa and strains of indigenous oral bacteria, both of which were isolated from the oral cavity of cystic fibrosis patients, was investigated by the phenomenon of the coaggregation reaction. A total of 22 strains of P. aeruginosa were isolated from the oral cavity of 17 patients and examined for their abilities to coaggregate with 5 strains each of Streptococcus sanguis, Streptococcus mitis, Actinomyces viscosus, and Actinomyces naeslundii. Coaggregation reactions were common between these oral bacteria and both the mucoid and nonmucoid variants of P. aeruginosa. All strains of P. aeruginosa were also able to agglutinate neuraminidase-treated or untreated human erythrocytes of blood types A, B, and O. Positive coaggregation reactions were further characterized by determining the effects of several sugars, and of heat and protease treatments of the bacteria. None of the coaggregtion reactions were inhibited by 0.05 M lactose, galactose, glucose, fucose, or mannose. All coaggregation reactions were dependent upon heat- and protease-sensitive components of the Pseudomonas. Thus, the interbacterial adhesions between P. aeruginosa and the oral bacteria studied appears to involve adhesins on the Pseudomonas cell, which bind to complementary receptors, on the cell surfaces of oral bacteria. The apparent prevalence and diversity of interbacterial adhesions between P. aeruginosa strains originating from the oral cavity of cystic fibrosis patients and strains of the indigenous oral bacteria suggest that some of these reactions may affect the extent to which P. aeruginosa colonizes in the oral cavity of cystic fibrosis patients, and thereby, influence susceptibility of the host to infection.     

Comparative acid tolerances and inhibitor sensitivities of isolated F-ATPases of oral lactic acid bacteria.

pH activity profiles and inhibitor sensitivities were compared for membrane ATPases isolated from three oral lactic acid bacteria, Lactobacillus casei ATCC 4646, Streptococcus mutans GS-5, and Streptococcus sanguis NCTC 10904, with, respectively, high, moderate, and low levels of acid tolerance. Membranes containing F1F0 ATPases were isolated by means of salt lysis of cells treated with muralytic enzymes. Membrane-free F1F0 complexes were then isolated from membranes by detergent extraction with Triton X-100 or octylglucoside. Finally, F1 complexes free of the proton-conducting F0 sector were obtained by washing membranes with buffers of low ionic strength. The pH activity profiles of the membrane-associated enzymes reflected the general acid tolerances of the organisms from which they were isolated; for example, pH optima were approximately 5.5, 6.0, and 7.0, respectively, for enzymes from L. casei, S. mutans, and S. sanguis. Roughly similar profiles were found for membrane-free F1F0 complexes, which were stabilized by phospholipids against loss of activity during storage. However, profiles for F1 enzymes were distinctly narrower, indicating that association with F0 and possibly other membrane components enhanced tolerance to both acid and alkaline media. All of the enzymes were found to have similar sensitivities to Al-F complexes, but only F1F0 enzymes were highly sensitive to dicyclohexylcarbodiimide. The procedures described for isolation of membrane-free F1F0 forms of the enzymes from oral lactic acid bacteria will be of use in future studies of the characteristics of the enzymes, especially in studies with liposomes.     

Comparative acid tolerances and inhibitor sensitivities of isolated F-ATPases of oral lactic acid bacteria.

pH activity profiles and inhibitor sensitivities were compared for membrane ATPases isolated from three oral lactic acid bacteria, Lactobacillus casei ATCC 4646, Streptococcus mutans GS-5, and Streptococcus sanguis NCTC 10904, with, respectively, high, moderate, and low levels of acid tolerance. Membranes containing F1F0 ATPases were isolated by means of salt lysis of cells treated with muralytic enzymes. Membrane-free F1F0 complexes were then isolated from membranes by detergent extraction with Triton X-100 or octylglucoside. Finally, F1 complexes free of the proton-conducting F0 sector were obtained by washing membranes with buffers of low ionic strength. The pH activity profiles of the membrane-associated enzymes reflected the general acid tolerances of the organisms from which they were isolated; for example, pH optima were approximately 5.5, 6.0, and 7.0, respectively, for enzymes from L. casei, S. mutans, and S. sanguis. Roughly similar profiles were found for membrane-free F1F0 complexes, which were stabilized by phospholipids against loss of activity during storage. However, profiles for F1 enzymes were distinctly narrower, indicating that association with F0 and possibly other membrane components enhanced tolerance to both acid and alkaline media. All of the enzymes were found to have similar sensitivities to Al-F complexes, but only F1F0 enzymes were highly sensitive to dicyclohexylcarbodiimide. The procedures described for isolation of membrane-free F1F0 forms of the enzymes from oral lactic acid bacteria will be of use in future studies of the characteristics of the enzymes, especially in studies with liposomes.     

Biosynthesis of phosphatidylcholine in bacteria

Phosphatidylcholine (PC) is the major membrane-forming phospholipid in eukaryotes and can be synthesized by either of two pathways, the methylation pathway or the CDP-choline pathway. Many prokaryotes lack PC, but it can be found in significant amounts in membranes of rather diverse bacteria and based on genomic data, we estimate that more than 10% of all bacteria possess PC. Enzymatic methylation of phosphatidylethanolamine via the methylation pathway was thought to be the only biosynthetic pathway to yield PC in bacteria. However, a choline-dependent pathway for PC biosynthesis has been discovered in Sinorhizobium meliloti. In this pathway, PC synthase, condenses choline directly with CDP-diacylglyceride to form PC in one step. A number of symbiotic (Rhizobium leguminosarum, Mesorhizobium loti) and pathogenic (Agrobacterium tumefaciens, Brucella melitensis, Pseudomonas aeruginosa, Borrelia burgdorferi and Legionella pneumophila) bacteria seem to possess the PC synthase pathway and we suggest that the respective eukaryotic host functions as the provider of choline for this pathway. Pathogens entering their hosts through epithelia (Streptococcus pneumoniae, Haemophilus influenzae) require phosphocholine substitutions on their cell surface components that are biosynthetically also derived from choline supplied by the host. However, the incorporation of choline in these latter cases proceeds via choline phosphate and CDP-choline as intermediates. The occurrence of two intermediates in prokaryotes usually found as intermediates in the eukaryotic CDP-choline pathway for PC biosynthesis raises the question whether some bacteria might form PC via a CDP-choline pathway.