Abolition of Swarming of Proteus by p-Nitrophenyl Glycerin: General Properties

Para-nitrophenyl glycerin (PNPG) was shown to be an effective agent to abolish the swarming of Proteus mirabilis and Proteus vulgaris on predried solid culture media. The level required to abolish swarming varied with the strain of Proteus, the components of the medium, and also with the conditions of incubation. Generally 0.1 to 0.2 mM PNPG effectively abolished swarming for at least 24 h with aerobic incubation. Levels of PNPG that abolished swarming showed no effect upon the growth of the cells, little or no effect upon the motility characteristics of the organisms, and no effect upon the cellular morphology. PNPG was found to be freely water soluble, stable to autoclaving, and to retain biological activity for at least one month in prepared culture media stored under refrigeration.     

Production of the therapeutic and prophylactic preparation enterobifidin on the basis of Bifidobacterium adolescentis MC-42

Production of Enterobifidin comprises preparation of culture media, reparation of lyophilized Bifidobacterium adolescentis MS-42 culture, preparation of starters, cultivation of bacteria in fermenters, biomass conservation, and its biological control. The preparation contains physiologically active bifidobacterium cells with high activities of growth (mu = 0.7 h-1, g = 1.0 h) and acid formation (titratable acidity is approximately 120-140 degrees T; acetate concentration, 0.50-0.75%; and lactate concentration, 0.33-0.50%). The antagonistic activity of these bacteria towards Escherichia coli 08, E. coli 086, E. coli 015, E. coli 0115, and E. coli 0101 amounts to 98.2;, to Proteus vulgaris 102, to 87.2; and Staphylococcus aureus 209p, to 83.2%. The bifidobacteria (with a titer of 10(9) CFU/ml) remained viable for two to five months.     

Search of promising strains of bifidobacteria and lactobacillus for the development of new biopreparations

For the first time the species composition of bifidobacteria and lactobacilli in clinically healthy young children has been studied. As revealed in this study, the dominating species of bifidobacteria are B. longum, B. adolescentis and B. infantis, while the dominating species of lactobacilli are Lactobacillus acidophilus and L. rhamnosus. In 83 isolated cultures of bifidobacteria and 34 isolated species of lactobacilli the activity of acid formation and the antagonistic activity with respect to Escherichia coli, Klebsiella pneumoniae, Staphylococcus aureus, Enterococcus faecalis, Clostridium perfringens have been tested. B. longum strain 58B, B. infantis strain 37B, L. rhamnosus strain 12L and L. acidophilus strain 27L, typical for children of this age group, having good antagonistic activity and pronounced acid-forming properties, have been selected. These strains hold good promise to be used as the basis for the development of a complex probiotic preparation for correcting intestinal microflora in young children.     

Biological characteristics of strains of lactobacilli, promising for use as eubiotics]

In the course of in vitro studies 3 Lactobacillus strains with pronounced antagonistic activity against some pathogenic and opportunistic bacteria (shigellae, enteropathogenic Escherichia, Proteus, staphylococci) were selected. In experiments on germ-free rats faint colonization by L. plantarum 37 was observed in the small intestine, as well as in the large intestine when low doses of these bacilli were introduced into the gastrointestinal tract of the animals. In vitro experiments demonstrated the decreased growth rate of this strain. The prophylactic administration of two eubiotic strains, L. plantarum 37 and L. fermentum 39, simultaneously with chloramphenicol to primates inhibited the growth of opportunistic bacteria, though L. fermentum 39 excessively suppressed the content of Escherichia coli in the enterobacterial population. The optimum biological effect was achieved with the therapeutic use of these three strains for the correction of dysbiotic disturbances caused by the administration of tetracycline in volunteers.     

The antagonistic activity of bifidobacteria in vitro and in vivo studied by using gnotobiological technology

The antagonistic activity of 4 strains of bifidobacteria (B. adolescentis 2 F1, B. longum Z4, B. breve R2 and B. bifidum G1), isolated from the vagina of healthy females of the reproductive age, with respect to Escherichia coli, Klebsiella ozaenae, Staphylococcus aureus, Enterococcus faecalis, Pseudomonas aeruginosa and Gardnerella vaginalis were studied in vitro and in vivo. The in vitro experiments revealed that all above-mentioned bifidobacteria were capable of inhibiting the growth of all indicator bacterial strains. Still of all the bifidobacteria under study had different levels of activity. B. adolescentis strain 2 F1 exhibited the highest inhibiting activity in vitro. In contrast to in vitro experiments, in vivo experiments with B. breve R2 demonstrated its high antagonistic activity with respect to E. coli. The data thus obtained indicate that in the study of antagonistic activity the use of the in vivo model as also expedient, for it is mainly in vivo that probiotic preparations show their activity.     

双歧杆菌对门静脉血内毒素影响初步研究

本实验以大鼠为动物模型,探讨在肠道微生态失调的状态下,用大量双歧杆菌(Bifidobacteria)灌服,借以拮抗外源性需氧革兰氏阴性杆菌的生长繁殖,从而使肠源性内毒素自门静脉的吸收减少。对正常鼠、脱污染鼠、再污染鼠、治疗鼠和非治疗鼠,进行不同肠段的双歧杆菌、需氧革兰氏阴性杆菌的测定和门静脉血内毒素测定。结果表明:双歧杆菌对肠道需氧革兰氏阴性杆菌的生长繁殖,具有明显的生物拮抗作用,同时伴有相应的门静脉血内毒素水平的降低。     

Structure of the O-polysaccharide of Proteus serogroup O34 containing 2-acetamido-2-deoxy-alpha-D-galactosyl phosphate

On mild acid degradation of the lipopolysaccharide of Proteus vulgaris O34, strain CCUG 4669, the O-polysaccharide was cleaved at a glycosyl-phosphate linkage that is present in the main chain. The resultant phosphorylated oligosaccharides and an alkali-treated lipopolysaccharide were studied by sugar and methylation analyses along with 1H and 13C NMR spectroscopy, and the following structure of the branched tetrasaccharide phosphate repeating unit of the O-polysaccharide was established: [carbohydrate structure: see text]The O-polysaccharide of Proteus mirabilis strain TG 276 was found to have the same structure and, based on the structural and serological data, this strain was proposed to be classified into the same Proteus serogroup O34.     

大口鲇致病菌的分离鉴定、系统发育分析及相关特性的研究

从四川鲶科鱼类养殖场濒死的大口鲇(Silurus meridionalis Chen)体内分离到3株优势菌,人工感染试验表明菌株TWN3致病性最强。通过形态学观察、生理生化测定,结合DNA的G Cmol%、16S rDNA序列同源性和系统发育分析,确定TWN3为普通变形杆菌(Proteus vulgaris)。再根据生理生化特性,可进一步确定TWN3属于普通变形杆菌(Proteus vulgaris)中的P.vulgarisBG3。该菌株不溶血;对庆大霉素等4种药物敏感;对小鼠有较强的致病性。生长特性研究表明,该菌最适生长温度37℃、最适pH6.0、37℃时盐度1.5%生长最适。普通变形杆菌致病性广泛,但作为养殖大口鲇的致病菌国内外未见报道。另外,对该菌的分类学研究、生长特性、感染途径及防治进行了讨论。     

Use of antibiotic-resistant Bifidobacterium in treating ulcerative-necrotic enterocolitis in young children

Ulceronecrotic colitis in premature children is accompanied by the development of pronounced intestinal dysbacteriosis characterized by a sharp increase in the number of Escherichia, enterococci, staphylococci and the appearance of opportunistic enterobacteria (Klebsiella, Serratia, Citrobacter, Proteus) in large amounts. Clinical convalescence was observed in 2 weeks in children receiving antibiotic-resistant bifidobacteria with antibiotics and in 3-4 weeks in children receiving commercial bifidobacteria with antibiotics, while children treated only with antibiotics showed no signs of clinical convalescence during the whole course of treatment. After the course of treatment the most effective recovery of the intestinal microflora was observed in the group of patients who had received antibiotics in combination with antibiotic-resistant bifidobacteria. It was manifested by a decrease in the number of Escherichia, Klebsiella, Proteus, Serratia, staphylococci and enterococci simultaneously with an increase in the number of endogenous lactobacilli and bifidobacteria. In those children who had received commercial bifidobacteria in combination with antibiotics or had been treated only with antibiotics the process of the recovery of the intestinal microflora was considerably less pronounced than in the patients of the first group; a decrease in the number of Escherichia, Klebsiella, Proteus and Serratia occurred to a lesser extent, and at the same time an essential increase in the number of enterococci and staphylococci was observed. The level of endogenous lactobacilli and bifidobacteria was considerably reduced. Antibiotic-resistant bifidobacteria actively took in the intestine of the patients. On day 5 after the course of treatment was over their level was (4.3 +/- 0.4) X 10(4) cells/g feces.     

Effect of polyvinylpyrrolidone on microbial cells

The oral administration of polyvinylpyrrolidone to rats produces no effect on the intestinal microflora. At the same time, polyvinylpyrrolidone solutions decrease the toxicity of Escherichia coli and Proteus vulgaris, produce a bactericidal and agglutinating effect.     

Evaluation of bifidobacteria for the production of antimicrobial compounds and assessment of performance in cottage cheese at refrigeration temperature

Twelve strains of bifidobacteria were identified which exhibited a broad spectrum of antagonistic activity against both Gram-positive and Gram-negative indicators, especially Pseudomonas species, using deferred antagonism spot plate assays. Inhibitory action was shown to be unrelated to hydrogen peroxide production and not solely dependent on acidity. However, attempts to detect inhibitory activity in cell-free supernatant fluids from these strains were unsuccessful. The production of inhibitory compound(s) by Bifidobacterium infantis NCFB 2255 was shown to be an unstable trait resulting in phenotypic alternation between production and non-production. Results from food trials using commercial cottage cheese which was inoculated with the inhibitor-producing strains of Bif. infantis NCFB 2255 and Bif. breve NCFB 2258 indicated that levels of Pseudomonas were reduced, but this observation was species-dependent. The viability of bifidobacteria themselves during storage in cottage cheese at refrigeration temperature was found to be a strain variable trait.     

利用培养组学技术分离鉴定体外降解胆固醇肠道细菌及其功能评价

高胆固醇是诱发心脑血管疾病的重要因素之一。目前国内外降低胆固醇的主要方式是药物治疗,但其存在费用高及副作用多的弊端。研究表明,肠道细菌对机体的胆固醇代谢有重要影响,但降胆固醇肠道细菌的筛选方式及功能评价却少有报道。本研究通过培养组学方法,使用牛胆汁酸或人工混合胆汁酸作为筛选条件,从健康人体肠道筛选出36种耐胆汁酸细菌。以鼠李糖乳杆菌GG株 (Lactobacillus rhamnosus GG,LGG) 作为阳性对照,设置0 g/L、0.3 g/L、3 g/L三种胆汁酸浓度组,对耐胆汁酸细菌进行体外降胆固醇能力评估,确定奇异变形菌、斯氏普罗威登斯菌、普通变形菌等10种细菌为降胆固醇优势菌。随后对其中6种降胆固醇优势菌——奇异变形菌、斯氏普罗威登斯菌、普通变形菌、潘氏变形菌、污蝇解克杆菌、雷氏普罗威登斯菌进行体外降甘油三酯能力评估以及人工胃液耐受能力评估。结果显示,上述6株细菌体外降甘油三酯能力均优于LGG。伴随人工胃液pH值的下降和作用时间的延长,6株细菌的生存率均下降。上述筛选实验及功能评价为进一步开发潜在的降胆固醇细菌制品提供研究基础。     

Morphology and Reproductive Processes of the L Forms of Bacteria II. Comparative Study of L Forms and Mycoplasma with the Electron Microscope

Representative electron micrographs, from the study of eight strains of L forms and one strain of Mycoplasma, are presented. A- and B-type L forms were derived from two strains of Proteus, two other L forms were derived from a diphtheroid and from a staphylococcus strain, and two strains (designated as LX) were isolated from L forms derived from a group A beta-hemolytic streptococcus and from a staphylococcus. The Mycoplasma strain was isolated from goats. Sections were made of young colonies grown within agar and from parts of surface colonies embedded in the agar. B-type L colonies of Proteus were produced by inoculation of bacteria into media containing penicillin. The large bodies developing from the bacteria and the organisms in B-type L colonies of Proteus, like the parent bacteria, had a cell wall consisting of a plasma membrane and an outer cell wall. The loss of rigidity in the cell wall indicated an alteration in its structure. The A-type L cultures of Proteus consisted of irregular branching masses extending in several directions, of small dense organisms corresponding to the elementary corpuscles present in cultures of Mycoplasma, and of intermediary forms. In contrast to the B-type, all organisms in the A-type colonies were surrounded by a single unit membrane corresponding to the plasma membrane of bacteria. The structures inside the cell membrane, both in the A- and B-type, seemed to correspond to the structure of the parent bacteria, which contained ribosomes and threads of DNA. The elementary corpuscles formed chains and filaments, and, apparently, these corpuscles took part in the multiplication by gradual enlargement. The organisms seen in the cultures of all L forms and Mycoplasma studied, except in the B-type L forms of Proteus, corresponded in size, shape, and structure, as well as in the development of elementary corpuscles, to the organisms in the A-type L form of Proteus. In contrast to the spherical organisms usually seen in broth cultures, the organisms in young cultures of Mycoplasma, which were grown within the agar, were similar in morphology, as well as in the discernible structure of the organisms, to L forms. Significant morphological and structural differences were not apparent between the L forms and Mycoplasma (in cultures grown within agar media) under the conditions of this investigation.     

Effects of Proteus vulgaris growth on the establishment of a cheese microbial community and on the production of volatile aroma compounds in a model cheese

Aims:  To investigate the impact of Proteus vulgaris growth on a multispecies ecosystem and on volatile aroma compound production during cheese ripening.
Methods and Results:  The microbial community dynamics and the production of volatile aroma compounds of a nine-species cheese ecosystem were compared with or without the presence of P. vulgaris in the initial inoculum. Proteus vulgaris was able to colonize the cheese surface and it was one of the dominant species, representing 37% of total isolates at the end of ripening with counts of 9·2 log₁₀ CFU g⁻¹. In the presence of P. vulgaris , counts of Arthrobacter arilaitensis , Brevibacterium aurantiacum and Hafnia alvei significantly decreased. Proteus vulgaris influenced the production of total volatile aroma compounds with branched-chain aldehydes and their corresponding alcohols being most abundant.
Conclusions:  Proteus vulgaris was able to successfully implant itself in a complex cheese ecosystem and significantly contributed to the organoleptic properties of cheese during ripening. This bacterium also interacted negatively with other bacteria in the ecosystem studied.
Significance and Impact of the Study:  This is the first time that the impact of a Gram-negative bacterium on cheese microbial ecology and functionality has been described.     

Structure of a new ribitol teichoic acid-like O-polysaccharide of a serologically separate Proteus vulgaris strain, TG 276-1, classified into a new Proteus serogroup O53

An unusual ribitol teichoic acid-like O-polysaccharide was isolated by mild acid degradation of the lipopolysaccharide from a previously non-classified Proteus vulgaris strain TG 276-1. Structural studies using chemical analyses and 2D (1)H and (13)C NMR spectroscopy showed that the polysaccharide is a zwitterionic polymer with a repeating unit containing 2-acetamido-4-amino-2,4,6-trideoxy-D-galactose (D-FucNAc4N) and two D-ribitol phosphate (D-Rib-ol-5-P) residues and having the following structure:[formula: see text] where the non-glycosylated ribitol residue is randomly mono-O-acetylated. Based on the unique O-polysaccharide structure and the finding that the strain studied is serologically separate among Proteus bacteria, we propose to classify P. vulgaris strain TG 276-1 into a new Proteus serogroup, O53.     

Effect of Streptococcus faecalis BIO-4R on intestinal flora of weanling piglets and calves

The effect of oral administration of Streptococcus faecalis BIO-4R, an antibiotic-resistant lactic acid bacterium, on the intestinal flora of weanling piglets and cows reared on antibiotic-containing diet was investigated. Fourteen days after administration of the bacteria, the intestinal flora of the piglets was examined. Animals of the administered group had stabilized lactic flora such as bifidobacteria, streptococci, and lactobacilli, whereas most animals of control group had reduced lactic flora. On the other hand, abundant yeasts were detected from the cecum, colon, and feces of the control animals, but the levels were significantly lower in the animals given strain BIO-4R. The density of Salmonella in the intestine appeared to be reduced after the administration of strain BIO-4R. The number of BIO-4R cells was shown to be 10 times lower in the duodenum and jejunum than in the ileum, suggesting that strain BIO-4R might have grown transiently in the ileum. The similar trend toward stabilization of the lactic flora was also observed in cows after administration of BIO-4R. In addition, an antagonistic effect of the strain against yeasts and Salmonella was suggested. These findings indicate that the oral administration of strain BIO-4R is one of the useful methods whereby the potentially deleterious effect of antibiotics on the intestinal flora of farm animals may be minimized.     

Effect of Streptococcus faecalis BIO-4R on intestinal flora of weanling piglets and calves.

The effect of oral administration of Streptococcus faecalis BIO-4R, an antibiotic-resistant lactic acid bacterium, on the intestinal flora of weanling piglets and cows reared on antibiotic-containing diet was investigated. Fourteen days after administration of the bacteria, the intestinal flora of the piglets was examined. Animals of the administered group had stabilized lactic flora such as bifidobacteria, streptococci, and lactobacilli, whereas most animals of control group had reduced lactic flora. On the other hand, abundant yeasts were detected from the cecum, colon, and feces of the control animals, but the levels were significantly lower in the animals given strain BIO-4R. The density of Salmonella in the intestine appeared to be reduced after the administration of strain BIO-4R. The number of BIO-4R cells was shown to be 10 times lower in the duodenum and jejunum than in the ileum, suggesting that strain BIO-4R might have grown transiently in the ileum. The similar trend toward stabilization of the lactic flora was also observed in cows after administration of BIO-4R. In addition, an antagonistic effect of the strain against yeasts and Salmonella was suggested. These findings indicate that the oral administration of strain BIO-4R is one of the useful methods whereby the potentially deleterious effect of antibiotics on the intestinal flora of farm animals may be minimized.     

乳粉中普通变形杆菌和奇异变形杆菌复合PCR-DHPLC检测技术的建立

应用复合PCR结合变性高效液相色谱(DHPLC)技术,建立乳粉中普通变形杆菌和奇异变形杆菌的高通量检测方法。根据普通变形杆菌的blaA和blaB基因及奇异变形杆菌的ureR基因序列分别设计特异性引物,复合PCR扩增的产物经DHPLC技术进行快速检测,并以37株参考菌株做特异性试验。试验结果表明,该方法具有很好的特异性,经复合PCR-DHPLC可同时检测乳粉中普通变形杆菌和奇异变形杆菌。该方法可以快速、准确、高通量检测普通变形杆菌和奇异变形杆菌,是乳粉中致病菌高通量检测的新技术。     

Investigation of hydrophobicity of Proteus vulgaris strains and ability of Proteus vulgaris and Proteus penneri strains to penetrate bladder membrane HCV T-29 cells

Proteus bacilli play a particularly important role in urinary tract infections (UTI). Fimbriae and adherence ability and hemolysins production (HpmA, HlyA) are one of the factors of pathogenicity of these bacteria. In this paper we describe the invasion of HCV T-29 transitional bladder urothelial cells carcinoma strains of P. penneri, as well as P. vulgaris strains belonging to different serogroups. The cytotoxic effect was observed at 8 hour of incubation of the tested cells with P. vulgaris O21 and the same effect (complete lysis) at 6 hours by P. vulgaris O4 (this strain manifests maximal activity in the production of HlyA hemolysin). P. penneri strains, produce different types of fimbriae, expressed similar bacterial invasiveness. The hydrophobic properties of 25 P. vulgaris strains were also tested and only 3 strains occur to have hydrophobic cell surface.     

Growth of salmonellas in different enrichment media

The usefulness of selenite-F (S-F), tetrathionate (MKT) and Rappaport-10 (R-10) broths as enrichment media to support growth of salmonellas either alone or in the presence of other competing organisms was studied. Their ability to support the growth of stressed salmonellas from water was also investigated. It was observed that R-10 was more inhibitory to competing organisms than MKT and S-F. It strongly inhibited the growth of Pseudomonas aeruginosa, Citrobacter freundii and Proteus vulgaris though not of Escherichia coli and Enterobacter aerogenes. It was more toxic, however, to small numbers of salmonellas than MKT and S-F. Tetrathionate was strongly inhibitory for E. coli and Ent. aerogenes but much less so for Proteus and Pseudomonas species. Selenite-F was much less inhibitory than MKT to Ps. aeruginosa and it did not inhibit growth of E. coli and Ent. aerogenes as much as MKT. Salmonellas were inhibited by all three enrichment media and none of them is ideally suited for direct use. Of the three media, R-10 was much more inhibitory to stressed organisms than S-F or MKT.