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PubMed1975

Biochemical basis of the serological cross-reactions between Brucella abortus and Yersinia enterocolitica serotype O:9.

A method based on the inhibition of agglutination is described that may be used for the differential serological diagnosis between B. abortus and Y. enterocolitica serotype O:9. An antigen with high immunological capacity was isolated from Brucella. This antigen inhibited both homologous and heterologous agglutination by Brucella antiserum, but only the heterologous agglutination by Yersinia antiserum. It proved to be constitued of a polysaccharide (N-acetylglucosamine, glucose, mannose and 2-keto-3-deoxyoctonic acid), a protein and a phosphoglycerid moiety. Lipid A was absent from the Brucella antigen. Incomplete polysaccharide synthesis of the Brucella antigen, with concomitant loss of serological specificity by the rough mutant has been described. Oligosaccharides containing N-acetylgalactosamine, glucose and galactose were isolated from the specific side chain of Yersinia lipopolysaccharide. Lipid A constituents were also identified in the latter.

Acetylglucosamine↗
PubMed1977

Bacteriophage-resistant mutants of Escherichia coli K12. Location of receptors within the lipopolysaccharide.

A series of mutants of Escherichia coli K12 resistant to lipopolysaccharide (LPS)-specific bacteriophages were isolated, and examined with regard to their general properties, phage typing, chemical analysis of their LPS, and genetic analysis. Fourteen classes of mutants were distinguished on the basis of phage typing and sensitivity to bile salts. Three of the mutant classes are sensitive to phages to which the parent is resistant. Mutants which are sensitive to bile salts generally lack heptose in their LPS, but two mutant classes are exceptions to this rule. Analyses of the sugars in the purified LPS of all mutant classes indicated that mutants were obtained which are blocked at most stages in core polysaccharide synthesis. On the basis of the chemical analysis, in conjunction with phage typing data and other known properties of the mutants, it is deduced which residue(s) is involved as a receptor for each of the phages used and which residues hinder these receptors. Some of the mutant classes do not seem to be changed in their LPS structure. Many of the mutations map in or near the rfa locus, but some are far removed from this region.

Bacteriophage Typing↗
PubMed1977

A study on fungal radioresistance and sensitivity.

Lower doses of gamma and X-rays (5 and 1 Krad, respectively) stimulated the spore germination and growth of 7 tested fungi. Higher doses (500 and 100 Krad, respectively) revealed that Epicoccum nigrum was the most radioresistant, while Alternaria humicola, Stemphylium verruculosum, Rhizopus nigricans, and Cladosporium herbarum were of moderate resistance. Aspergillus niger and Penicillium oxalicum were the most radiosensitive. Protein and polysaccharide synthesis were inhibited in the mycelium developed from irradiated spores; the inhibition was a function of dose. Study of the different factors (morphological, biological, and biochemical) indicates that variability in fungal resistance to ionizing radiation is, most probably, an inherent character, connected with mycelial water content and the natural production of a chemical substance(s) that acts as a radioprotector(s).

Amino Acids↗
PubMed2026

Phage vB_KpnM_NB cocktail synergizing with amikacin in inhibiting persister cells of Klebsiella pneumoniae.

UNLABELLED: The emergence of multidrug-resistant Klebsiella pneumoniae (KPN) and antibiotic-tolerant persister cells poses a significant challenge to existing anti-infection therapies. Given the urgent need for sustainable alternatives to antibiotics, phage cocktails are emerging as a promising alternative to control K. pneumoniae infections. We isolated three lytic phages vB_KpnM_NB (1-3) from Ningbo environmental samples, classified them into the Drexlerviridae family, and determined the biological characteristics of two representative phages. Genomic analysis confirmed that these phages are closely related and lack resistance and virulence genes, ensuring biosafety. Subsequently, a stable KPN persister model was established using amikacin, with a biphasic killing pattern observed during treatment. At a multiplicity of infection of 10, the phage cocktail eliminated 99.00% of persister cells, while individual phages were less effective. The phage cocktail also inhibited persister-derived biofilm formation, showing improved results when combined with amikacin. This combination significantly reduced capsule polysaccharide production in persisters, weakening the outer membrane barrier. These findings demonstrate that the phage cocktail-amikacin combination effectively targets planktonic cells, persister cells, and biofilms, providing a promising strategy against persisters and recurrent K. pneumoniae infections. IMPORTANCE: This study fills the critical gap in understanding how phage cocktails synergize with amikacin against K. pneumoniae persister cells. By constructing a highly specific phage vB_KpnM_NB cocktail, establishing a stable persister model, and performing in vitro bactericidal and biofilm assays, we demonstrate that the cocktail effectively eliminates planktonic cells, persisters, and biofilms. We clarify the core synergistic mechanism: inhibiting capsular polysaccharide synthesis, improving phage adsorption, and disrupting the bacterial outer membrane barrier. These findings provide experimental evidence for the prevention and control of multidrug-resistant and carbapenem-resistant K. pneumoniae persister infections, establishing a safe and effective phage-antibiotic combination therapy. The results are crucial for addressing antibiotic tolerance and controlling chronic, recurrent infections. They hold significant theoretical and translational value for the treatment of refractory infections in clinical settings and offer new insights into the development of novel antimicrobial strategies.

Klebsiella pneumoniae↗
PubMed1977

Identification, separation, and preliminary characterization of invertase and beta-galactosidase in Actinomyces viscosus.

The initial step of disaccharide dissimilation by Actinomyces viscosus serotype 2 strain M-100 was studied. Sucrase activity was found in the 3,000 X g particulate fraction and the 37,000 X g soluble fraction of the cells, whereas lactase activity was found almost exclusively in the 37,000 X g soluble fraction. Neither sucrase nor lactase activity was appreciable in the culture liquor. Sucrose phosphorylase, alpha-glucosidase, and polysaccharide synthesis activities were not observed in the soluble cell fraction. The sucrase was identified as invertase (EC 3.2.1.26; beta-D-fructofuranoside fructohydrolase). The lactase was identified as beta-galactosidase (EC 3.2.1.23; beta-D-galactoside galactohydrolase). The enzymes in the 37,000 X g soluble fraction were separable by diethylamino-ethyl-cellulose chromatography, giving one beta-galactosidase peak and one major and one minor invertase peak. Acrylamide gel electrophoresis showed different electrophoretic mobilities of the enzymes. The molecular weight of the beta-galactosidase is about 4.2 X 10(5) and that of invertase is about 8.6 X 10(4). The beta-galactosidase has a Km for lactose of about 6 mM and a pH optimum between pH 6.0 and 6.5. The major invertase component has a Km for sucrose of about 71 mM and a pH optimum between pH 5.8 and 6.3.

Actinomyces↗
PubMed2025

Accurate serotype identification of Streptococcus pneumoniae using nanopore Cas9-targeted serotype identification (nCATSerotyping).

Streptococcus pneumoniae (pneumococcus) is a leading cause of community-acquired pneumonia and invasive diseases, particularly among children and the elderly. The introduction of pneumococcal conjugate vaccines has significantly reduced invasive pneumococcal disease, but the prevalence of non-vaccine serotypes and newly emerging serotypes is increasing globally. Thus, accurate serotyping is essential for epidemiological surveillance and the development of next-generation multivalent pneumococcal vaccines. Conventional serotyping methods, including multiplex polymerase chain reaction (mPCR), monoclonal antibody (mAb) assays, and Quellung reaction using rabbit antisera, are limited by serotype coverage and cross-reactivity, making the detection of new or emerging serotypes challenging. In this study, we developed a nanopore Cas9-targeted serotyping (nCATSerotyping) platform, which employs Cas9-mediated enrichment of the capsular polysaccharide synthesis locus followed by Oxford Nanopore sequencing. Applying this method to 276 clinical pneumococcal isolates collected in South Korea (2018-2020), we achieved a serotyping success rate of 97.10% (268/276), significantly outperforming conventional methods such as mAb and mPCR, which identified only 76.45% (211/276) of isolates. Whole-genome sequencing of the remaining eight non-typeable isolates revealed them to be non-pneumococcal (oral streptococci), confirming 100% accuracy for S. pneumoniae serotyping. Importantly, our method identified emerging and underrepresented serotypes, including serotype 13 and null capsule clade strains. nCATSerotyping offers a rapid, accurate, and comprehensive solution for pneumococcal serotyping, with significant advantages in identifying novel and non-typeable strains. This scalable platform will be a valuable tool for global serotype surveillance and next-generation multivalent pneumococcal vaccine development.IMPORTANCEAccurate pneumococcal serotyping is critical for vaccine development and epidemiological surveillance, particularly as non-vaccine serotypes emerge following widespread pneumococcal conjugate vaccine implementation. Current serotyping methods face significant limitations in coverage and accuracy, identifying around 76% of pneumococcal isolates and failing to detect emerging serotypes like serotype 13 and null capsule clades. The nanopore Cas9-targeted serotyping platform addresses these critical gaps by achieving 100% serotyping accuracy for confirmed Streptococcus pneumoniae isolates while identifying previously undetectable strains that conventional methods missed. This comprehensive approach is essential for monitoring vaccine effectiveness, understanding serotype replacement patterns, and informing next-generation vaccine development strategies. Furthermore, the identification of misclassified oral streptococci highlights the diagnostic precision needed for accurate pneumococcal surveillance, ensuring that epidemiological data accurately reflect true pneumococcal disease burden and serotype distribution patterns.

Streptococcus pneumoniae↗
PubMed2026

By comparing the effects of Lactobacillus paracasei KL1 and BK56 strains on yogurt quality, the optimal consumption time for 2 compound fermented yogurts was determined.

This study investigated the effects of 2 Lactobacillus paracasei strains, KL1 and BK56, on the physicochemical properties, microstructure, texture characteristics, and sensory quality of a compound fermented yogurt system (GK107: L. paracasei KL1, Leuconostoc mesenteroides G12S, Chr. Hansen Commercial Starter Culture; G56107: L. paracasei BK56, L. mesenteroides G12S, Chr. Hansen Commercial Starter Culture), and further integrated genomic and metabolomic analyses to infer their shelf-life and optimal consumption period. The results showed that GK107 yogurt maintained stable quality throughout the 28-d storage period (at d 28: pH 4.07; titratable acidity 93.25 °T; exopolysaccharide content 0.31 g/L; water-holding capacity 53.05%; sensory score 83), and rapidly formed a stable gel structure that persisted for an extended duration. In contrast, the quality of G56107 yogurt deteriorated during the later stage of storage (at d 28: pH 4.0; titratable acidity 98.4 °T; exopolysaccharide content 0.31 g/L; water-holding capacity 51.3%; sensory score 67). Genomic analysis revealed that, compared with the L. paracasei KL1 strain, the L. paracasei BK56 strain carried loss-of-function mutations in multiple key genes associated with flavor synthesis, polysaccharide metabolism, and proteolysis, including alsS, prtP, glpO, AWC33_RS01450, AWC33_RS00855, AWC33_RS01070, and AWC33_RS01805. These mutations may have played a role in the gradual flavor deterioration, and weak post-acidification control observed in G56107 yogurt during prolonged storage. Based on the above results, it is reasonable to suggest that GK107 yogurt is suitable for long-term storage with an optimal consumption period of 14 to 28 d, whereas G56107 yogurt is more suitable for short-term storage and recommended for consumption within the first 14 d.

Genomics↗
PubMed1976

The butanol-soluble proteins of Klebsiella aerogenes.

Membranes of Klebsiella aerogenes were extracted with acid butanol at pH 4.2 and the solubilized proteins were fractionated by precipitation at 0 degrees C and -20 degrees C. The -20 degrees C precipitate was examined by polyacrylamide gel electrophoresis in sodium dodecyl sulphate. Several enzyme activities associated with polysaccharide biosynthesis have been identified in the crude -20 degrees C precipitate and an attempt was made to separate the enzymes from other protein. The role of certain of these enzymes in a control mechanism for polysaccharide synthesis in K. aerogenes has been postulated.

Amino Acids↗
PubMed1977

Experimental arthritis in rabbit knees: a study of relief of pressure on one tibial plateau in immature and mature rabbits.

The Hall model of experimental arthritis of the knee (excision of the lateral femoral condyle) offers a unique way of investigating the earliest changes of degenerative arthritis. Predictable degeneration of the lateral tibial plateau occurs when there is lack of contact with an opposing surface. Previous work by Hall and Thompson has not concentrated upon the early changes in proven mature animals of optimum age for a study of simulated osteoarthritis. In this work using mature rabbits, two important findings are documented. The earliest observed change in the joint, beginning at 7 days is loss of matrix protein-polysaccharides from the lateral plateau, with no evidence of change in the subchondral bone until much later; in this respect mature differ from immature animals. Furthermore, there is strong evidence for late (three to 6 months) protein-polysaccharide synthesis in the previously degeneraing articular cartilage in which the residual cells reconstitute an effective matrix.

Age Factors↗
PubMed1977

Genetic determinants of the synthesis of the polysaccharide capsular antigen K27(A) of Escherichia coli.

Most of the his+ hybrids from crosses between the Escherichia coli donor Hfr45(O8:K27) and different E. coli O9 recipients expressed the donor O8 antigen specificity and produced the capsular antigen K27. Therefore these hybrids must have inherited the his-linked donor rfb region determining the synthesis of O8- specific polysaccharides as well as his-linked genes involved in K27 antigen synthesis. In the living state these hybrids were inagglutinable in O8 antiserum like the donor cells. However, when E. coli K12 and O8:K42- were used as recipients most of the his+ hybrids were agglutinable in O8 and K27 antisera. The amounts of K27 antigen present in these hybrids, designated as K27i (intermediate) forms, were sufficient to evoke the production of K27 antibodies in rabbits, but insufficient to inhibit O-agglutination of the respective cells. The additional transfer of the trp region of E. coli O8:K27 into such K27i forms frequently resulted in O-inagglutinable K27+ hybrids. This is attributed to the introduction of trp-linked genes which apparently play a role in the synthesis of K27 capsular antigen. Tus it is concluded that at least two gene loci, one close to his and the other close to trp, are required for the synthesis of the complete capsular antigen K27.

Antigens, Bacterial↗
PubMedDate not supplied

[More active synthesis of the polysaccharide, pullulan, by polyploid cultures of Pullularia pullulans].

The polyploid strains of Pullularia pullulans differ from each other and from the haploid strain in the amount of the polysaccharide pullulan liberated into the cultural broth. The highest pullulan synthesizing activity (per unit of the assimilated carbon source and the synthesized biomass) was manifested by the diploid strain of P. pullulans 1125(13) whose cells produced more pullulan (by 75%) within three days of growth than the cells of the haploid culture. The content of pullulan calculated per unit area of the cell surface increased with the level of ploidy: 1.5-1.8 times in the diploid cultures and 3.4 times in the tetraploid culture cf. the parent haploid culture. Apparently, the polyploidy of the P. pullulans culture was accompanied with mutations involved in the synthesis of the extracellular polysaccharide.

Culture Media↗
PubMedDate not supplied

Acrylamide gel electrophoretic studies of extracellular sucrose-metabolizing enzymes of Streptococcus mutans.

This study explored the use of acrylamide gel electrophoretic methods to determine the numbers and types of extracellular sucrose-metabolizing enzymes produced by particular strains of S mutans. Strains HS-6, SL-1, FA-1, and NCTC 10449 were cultured in a chemically defined medium and the extracellular proteins elaborated by the organisms were isolated and subjected to acrylamide gel electrophoresis. Patterns of protein components and sucrose-metabolizing enzymes were then delineated. Three types of sucrose-metabolizing enzymes were observed. One type was involved in the synthesis of polysaccharides that were insolubilized in the gels. Another type was involved in the synthesis of water-soluble polysaccharides. A third type was involved in the splitting of sucrose into reducing sugars without polysaccharide synthesis. Each pattern was distinctive with regard to the numbers, proportions, and types of enzyme components present and their migratory characteristics. From two to at least six components were observed amont the enzyme activity patterns per strain. Extracellular protein patterns showed from 12 to 20 components per strain. Comparative data on growth in the chemically defined medium and Todd-Hewitt broth were also presented. Better growth levels were obtained in all instances with the chemically defined medium over comparable periods of time.

Culture Media↗
PubMed1977

Synthesis of "group polysaccharide" by membranes from Streptococcus pyogenes and its stabilized L-form.

Rhamnose and N-acetylglucosamine (GlcNAc) are incorporated from thymidine 5'-diphosphorhamnose and uridine 5-diphospho-N-acetylglucosamine (UDPGlcNAc) into membrane fragments prepared from Streptococcus pyogenes but not into membrane fragements prepared from a stabilized L-form of this organism. Incorporation from TDPrhamnose is partially dependent upon UDPGlcNAc and vice versa. The oligomeric GlcNAc and rhamonose-containing products are easily extracted from membrane particles by sedimentation through detergent solutions. They are substantially extracted into methanol but not into chloroform-methanol (2:1). When product containing both radioactive rhamnose and GlcNAc is deacetylated and hydrolyzed briefy in acid, glucosaminyl rhamnose is obtained, byt not higher oligomers, suggesting that oligomer synthesis in vitro is terminated because unidentified wnzymatic requirements are not satisfied. The data are consistent with the assembly of group A-specific polysaccharide at the cellular membrane with participation of a lipoid anchor (acceptor) molecule.

Acetylglucosamine↗
PubMed1977

[Synthesis of extracellular polysaccharides by various forms of Streptococcus salivarius].

Streptococcus salivarius synthesizes extracellular polysaccharides (EPS) from saccharose under aerobic and anaerobic conditions. Under aerobic conditions primarily soluble fructanes are formed. There are no differences between the EPS synthesized by r- and s-forms under aerobic conditions; anaerobically the s-forms produce more EPS and more fructane than the r-forms. As compared to streptococcus mutans, the streptococcus salivarius produces considerably more fructane and insoluble glucane under the conditions used.

Aerobiosis↗
PubMed1976

Abiotic synthesis of some polysaccharide-like and polypeptide-like structures in cold plasma.

Starting from methane, ammonia and water vapors polypeptide-like and polysaccharide-like structures have been synthesized in cold plasma conditions. By gel filtration analysis of the raw product eight fractions of different molecular weight have been separated. The partial chemical nature of each fraction and of the hydrolysis products has been determined using elementary analysis, IR spectroscopy, paper, thin-layer and gas-chromatography. It is noteworthy that higher proportions of H2O in the feed composition increase the amount of sugar and polysaccharide-like compounds in the reaction product.

Amino Acids↗
PubMed1977

Mycobacterium smegmatis fatty acid synthetase. Polysaccharide stimulation of the rate-limiting step.

An initial activity burst lasting 5 to 10 s is observed for both de novo synthesis with acetyl-CoA as primer and for elongation of palmitoyl-CoA catalyzed by the multienzyme complex fatty acid synthetase from Mycobacterium smegmatis. After the initial burst, synthetase activity slows at least 6-fold to the steady state rate. The size of the initial burst is proportional to the amount of synthetase protein and corresponds to the synthesis of a small number C three to five) of C24 or C26 acyl chains per mol of enzyme. During the initial burst, C24, C26 acyl enzyme is formed and can be isolated by ammonium sulfate precipitation. On incubation with CoA, enzyme-bound acyl chains undergo transacylation to form the corresponding CoA derivatives. Diffusion of C24-CoA and C26-CoA from the enzyme is slow and rate-limiting for overall fatty acid synthesis. Mycobacterial polysaccharides markedly accelerate this rate-determining step but bovine serum albumin does not. This facilitation of product diffusion accounts for the large stimulation of de novo synthesis and of elongation of mycobacterial polysaccharide. It is also shown that the high apparent Km for acetyl-CoA (approximately 400 micrometer) in the steady state reflects the substrate concentration required to shift the product pattern in favor of shorter chain fatty acids (C16,C18). These conditions circumvent the slow, rate-limiting diffusion of C24-CoA and C26-CoA.

Acetyl Coenzyme A↗