Penicillin-binding proteins and carboxypeptidase/transpeptidase activities in Proteus vulgaris P18 and its penicillin-induced stable L-forms

The originally penicillin-induced, wall-less stable L-forms of Proteus vulgaris P18, isolated by Tulasne in 1949 and since then cultured in he absence of penicillin, have kept the ability to synthesize the seven penicillin-binding proteins and the various DD- and LD-peptidase activities found in the parental bacteria and known to be involved in wall peptidoglycan metabolism. The stable L-forms, however, secrete during growth both the highly penicillin-sensitive, DD-carboxy-peptidase-transpeptidase penicillin-binding protein PBP4 (which in normal bacteria is relatively loosely bound to the plasma membrane) and the penicillin-insensitive LD-carboxypeptidase (which in normal bacteria is located in the periplasmic region).     

Alkenyl and alkyl phosphatides in Proteus and its L forms

Summary Alkenyl and alkyl phosphatides were characterized in the lipids of Proteus mirabilis, grown in aerobic conditions. These classes of lipids are also present in the stable L-forms derived from Proteus. The amounts of alkenyl phosphatides are very low, compared to the other bacteria which contain this lipid. For alkyl phosphatides, the amounts are of the same order as that found in anaerobic bacteria.     

Polyacrylamide Gel Identification of Bacterial L-Forms and Mycoplasma Species of Human Origin

Polyacrylamide gel electrophoretic patterns of acidified phenol extracts prepared from whole cells can be used for the identification of bacterial L-forms and Mycoplasma species of human origin. Ten human Mycoplasma serotypes and eight L-forms belonging to five different genera were studied. The gel patterns were sufficiently distinct and reproducible that it was possible not only to identify L-forms at the genus level (group with streptococci) and different Mycoplasma serotypes but also to differentiate between the two of them. The parentage of L-forms of Streptobacillus moniliformis L1, Listeria monocytogenes, Streptococcus MG, and Staphylococcus aureus Smith strain was established by relating their gel patterns directly to parent bacteria. It was found that an L-form designated S. moniliformis An (ATCC 14220) was actually an L-form of Proteus. In addition, it was shown electrophoretically that no relationship existed between the Streptococcus MG L-form and M. pneumoniae. The applicability of this method as a diagnostic and taxonomic tool for the differentiation of L-forms and mycoplasmas is discussed.     

Generic Identification of L-Forms by Polyacrylamide Gel Electrophoretic Comparison of Extracts from Parent Strains and Their Derived L-Forms

A polyacrylamide gel electrophoretic method was used for identification of L-forms in the genera Streptococcus, Staphylococcus, and Proteus, by comparing phenol-acetic acid-water extracts of homologous parent L-form pairs to one another and to other pairs. The method requires only milligram quantities of material for analysis. The standard patterns for the strains used in this study are shown as pictures of the gels and as densitometric tracings of appropriate gels. They show that, despite occasional minor differences in some organisms, the gel electrophoretic patterns of homologous L-forms and bacterial pairs are sufficiently similar-as well as sufficiently dissimilar from patterns of other genera-to permit generic identification of an unknown L-form by reference to patterns derived from possible parental bacteria.     

Fatty acid composition of membranes of L-forms ofStreptococcus faecalis andProteus mirabilis cultured at different osmolalities

The fatty acid composition of membranes of L-forms ofStreptococcus faecalis andProteus mirabilis cultured at different osmolalities and in different osmotic stabilizers was examined.S. faecalis L-forms cultured with sucrose in the medium showed a decrease in the unsaturated fatty acid C₁₈₁ and an increase in the C₁₈ fatty acid and C₁₉ cyclopropane fatty acid. Fatty acid composition ofS. faecalis L-forms cultured in medium containing 1.8% NaCl was similar to the fatty acid composition of L-forms cultured in brain-heart infusion broth (BHI) without osmotic stabilizer and was between the composition of fatty acids of L-forms in BHI with sucrose and that in BHI without 0.5 M sucrose. InProteus mirabilis L-forms, there were differences between L-forms cultured with and without sucrose, but these differences were not comparable to the changes observed inS. faecalis L-forms.P. mirabilis L-forms cultured with and without NaCl in the medium had similar fatty acid compositions.     

Chemotactic activity of L-forms and mycoplasma.

L-forms of Streptococcus faecalis, Escherichia coli and Proteus mirabilis showed significantly less chemotactic activity for normal human leucocytes than did parent bacterial forms which were strongly chemotactic. Mycoplasma pneumoniae and Mycoplasma hominis did not demonstrate chemotactic activity.     

Chemical, Biological, and Structural Properties of Stable Proteus L Forms and Their Parent Bacteria

Proteus L forms were disrupted by osmotic shock, and the sedimentable material present in the homogenate was further fragmented in a Sorvall pressure cell. The pressure cell was also used for disrupting normal Proteus cells. The homogenates obtained were fractionated by differential centrifugation. Purified endotoxins were isolated from the major fractions by phenol extraction. Material extracted with phenol from the membrane fraction of the L forms was about as toxic and pyrogenic on a weight basis as the typical enterobacterial endotoxins isolated from cell walls of normal bacteria. The yield of extract from L forms was about one-third of that from an equal weight of normal bacteria. No differences in the gross chemical composition of the phenol extracts from the L forms and the normal cells could be ascertained. A close serological relationship existed between extracts obtained from two L forms and their respective parent bacteria, but no such relationship was found in the case of the third L form studied and its parent bacterium. Diaminopimelic acid was not detected in the membranes of the L forms, but these membranes contained most of the succinic dehydrogenase of the organisms. Only small amounts of this enzyme were present in the wall fraction of normal bacteria. The data obtained suggest that precursors of the Proteus endotoxins are formed either in the soluble protoplasm of normal cells and L forms or at sites on the membrane from which they are readily liberated into the protoplasm, whereas the final steps of the synthesis of these toxins take place at the cytoplasmic membrane. In normal cells, much endotoxin is transported to and concentrated in the walls.     

L-forms ofHaemophilus influenzae: Growth and reversion as influenced by a strain ofNeisseria perflava

Factors excreted into the medium by various bacteria and fungi exert a significant effect on the growth and revertibility of bacterial L-forms. Studies with L-forms ofHaemophilus influenzae have been facilitated by a strain ofNeisseria perflava, which not only promotes the growth of the L-forms on media of normal osmolarity (horse blood agar), but also enhances the revertibility of the L-forms to the parent bacterium. L-forms derived fromN. perflava also exhibit this growth-enhacing property, an effect that does not appear to be related to X and V factors. Preliminary characterization of the active component produced byNeisseria perflava indicates that it is present in broth filtrate, diffusible through agar, heat labile, and of large molecular size.     

Antimicrobial activities of daunorubicin and adriamycin derivatives on bacterial and protoplast type L-form cells of Bacillus subtilis 170, Escherichia coli B, and Proteus mirabilis VI. Structure--activity relationship

The antibacterial activity of ten N-alkylated derivatives of daunorubicin and adriamycin as well as of 5-iminodaunorubicin has been tested by using Bacillus subtilis 170, Escherichia coli B, and Proteus mirabilis VI and their stable protoplast type L-forms in an agar diffusion test. Eight of the substances showed similar activities against B. subtilis and the L-forms of all test organisms, but no activity against the bacterial forms of E. coli and P. mirabilis. The cell wall of these gram-negative bacteria is responsible for this resistance by not allowing the antibiotics to enter the cells. The piperidino compound N-(CH2)5 daunorubicin shows 2-4 times higher activity against B. subtilis and all L-forms in comparison to daunorubicin and the other derivatives. Five of the substances were inactive against all test strains. Their inactivity seems to be associated with the larger substituents at the C-3' position. Relations between molecular structure and activity are discussed considering data about the interaction with DNA and the antitumor activity. Stable protoplast type L-forms and their bacterial forms represent a suitable and effective test system to screen for more effective substances and to get more information about their mode of action.     

p53、p16和细菌L型DNA在卵巢肿瘤中的表达及意义

目的 探讨p53、p16抑癌基因和细菌L型DNA在卵巢肿瘤中的表达及临床意义。方法 用原位核酸杂交方法检测了101例卵巢肿瘤中的p53、p16抑癌基因以及L型菌DNA的阳性信号。结果 恶性肿瘤中p53原位核酸杂交mRNA的阳性信号,恶性肿瘤明显高于良性肿瘤（P＜0.01）,p16原位核酸杂交mRNA的阳性信号,恶性肿瘤则低于良性肿瘤（P＜0.01）,2种抑癌基因的指标与肿瘤的病理分级及患者预后有明显相关性（P＜0.05）。而细菌L型的阳性信号与病理分级、有腹水比无腹水者、腹腔淋巴结有转移比无转移者差异有显著性（P＜0.05）。结论 p53 mRNA在卵巢肿瘤中有不同程度异常表达,p16 mRNA表达与肿瘤细胞分化呈负相关,2种抑癌基因均可作为判断卵巢肿瘤生物学行为及患者预后参考指标,L型感染有可能成为诱发肿瘤因素之一,它们与抑癌基因可能有协同致瘤作用。研究细菌L型感染与肿瘤的关系,具有重要的临床应用价值。     

An ELISA for the detection of Bacillus subtilis L-form bacteria confirms their symbiosis in strawberry

AIMS: To develop an ELISA for the detection of antigens derived from stable Bacillus subtilis L-form bacteria and to detect these in plants injected with L-form bacteria. METHODS AND RESULTS: A sandwich ELISA was developed and its specificity was investigated using L-forms and cell-walled forms of B. subtilis, different Bacillus species and a range of bacteria isolated from glasshouse-grown strawberry plants. The detection limits of the ELISA were approximately 10(3) viable cells ml(-1) for L-forms compared with 10(7) viable cells ml(-1) for cell-walled forms. Results showed that L-forms survived and moved within strawberry tissues injected with L-form bacteria. CONCLUSION: An ELISA that selectively detects B. subtilis L-form bacteria was developed and shown to confirm the presence of L-forms in plants. SIGNIFICANCE AND IMPACT OF THE STUDY: This will be a valuable rapid method to further studies on L-form plant interactions.     

慢性肾盂肾炎L型细菌感染及耐药分析

目的了解L型细菌在慢性肾盂肾炎的感染及耐药状况。方法对71例患者清洁中段尿做普通细菌培养(B型)、L型细菌培养(L型)及耐药分析。结果细菌阳性率为77.5%,其中单独L型阳性率为49.3%、B型与L型混合感染为15.5%,而B型阳性率仅为12.7%。主要是大肠埃希菌,其次是葡萄球菌;青霉素及头孢噻肟均有较高的耐药率(88.9%及73.6%)。结论L型细菌在慢性肾盂肾炎感染中占主导,β-内酰胺类药物有较高的耐药性,临床治疗应据药敏结果合理选择及时调整抗生素。     

Novel shuttle vectors for improved streptokinase expression in streptococci and bacterial L-forms

Novel shuttle vectors of small size and increased copy number capable of replication in Escherichia coli, L-forms of Proteus mirabilis, and streptococci were constructed from a streptococcal erythromycin-resistant plasmid and an Escherichia coli phasmid. The streptokinase gene, skc, was inserted into one of them, and skc expression was studied in Streptococcus sanguis, Streptococcus lactis, and in an L-form strain (LVI) of Proteus mirabilis. The new streptokinase shuttle plasmid, pMLS10 (7.3 kb), specified higher Skc yields in all hosts when compared to pSM752 constructed previously. In particular Proteus mirabilis LVI(pMLS10) proved to be the most productive host, exhibiting complete secretion of the active protein at yields as high as 24000 unit per ml.     

Localization of enterobacterial common antigen: Proteus mirabilis and its various L-forms.

An investigation of Proteus mirabilis wild-type strains and their various derived L-forms shows that the enterobacterial common antigen (ECA) is localized in the outer membrane of the cell envelope of these strains. In strains where the outer membrane is lacking (stable protoplast L-forms) or where its amount is reduced (spheroplast UL19) no ECA or only reduced amounts of it are detected by serological tests or by ferritin-labeling techniques.     

细胞壁缺陷细菌生物氧化特性的观察

The L-forms were induced from Staphylococcus aureus,Escherichia coli and Bacdtos cereus by β-lactam anhbiohcs and then observahons on the proPenies of oxygen requlrement sugar fermentahon and sensihve tO cyanide of the Lforms were done. The resultS were shown that the Lforms derived from the obligate aerobe or the faCultative anaerobe did not ferment sugars and were highly oxygendePendent and more sensihve tD cyedde than their Parent bacteria The metabolic achvihes which were same as the Parent bacteria of …     

Induction, growth and antibiotic production of Streptomyces viridifaciens L-form bacteria

AIMS: To induce, cultivate and investigate the characteristics of L-form bacteria derived from the filamentous actinomycete Streptomyces viridifaciens. METHODS AND RESULTS: L-forms were induced in a liquid medium supplemented with lysozyme and penicillin. A stable culture which no longer required inducing agents but could still revert, was obtained by the twelfth subculture. The specific growth rate of stable L-forms was faster (0.751) than unstable L-forms (0.361). After the exponential growth phase, the cell diameter continued to increase, as did the percentage of vacuoles. Morphologically, the L-forms appeared as spherical bodies with no signs of differentiation and were sensitive to osmotic stress, indicating removal of the cell wall. The L-forms produced secondary metabolites although much lower levels of antibiotic were assayed in the L-forms compared with the cell walled forms. CONCLUSION: Stable L-form bacteria were induced from S. viridifaciens and their growth characterized. The L-forms produced secondary metabolites. SIGNIFICANCE AND IMPACT OF THE STUDY: Stable Streptomyces L-forms were induced and have potential as biocontrol agents.     

Isolation of L-Forms in a Clinical Microbiology Laboratory

Previous studies have demonstrated that L-forms of bacteria may play a role in persistent, chronic, or recurrent urinary-tract infections. A 2-year program was initiated to determine the feasibility of culturing for L-forms on a routine basis, and to determine the effectiveness of such a program. In relation to the total number of specimens, few L-forms were actually isolated. In comparison with the amount of equipment and technician time required, the return was negligible; only 0.5% of all urine specimens were positive for L-forms. An increase to only 1.2% was noted when culturing for L-forms was limited to patients with a diagnosis of bacteriuria or pyelonephritis. It is recommended that this technique be reserved for those patients with a long history of recurrent urinary-tract infections, after other attempts to cure the patient have met with failure.     

Processes of bacterial L transformation and L form reversion in the body of argasid ticks

The experimental infection of tampan ticks (Ornithodoros moubata) with the bacterial cultures of Listeria monocytogenes and Salmonella typhimurium, as well as with their L-forms, was carried out. These experiments demonstrated that both the L-transformation of bacteria and the reversion of their L-forms into the initial bacterial culture could occur in the body of the ticks.     

L-forms of bacteria in the urine of patients with chronic pyelonephritis]

The authors present the data concerning the study of 200 patients suffering from chronic pyelonephritis; in 28 of these L-forms of bacteria were revealed in the urine. Of 46 L-cultures isolated from these patients 13 reversed into bacterial forms, 8 failed to reverse and were referred to the stable L-forms; the rest 25 L-cultures perished during the 8th--10th passage. This led to a supposition that the relapses and exacerbations of the infectious process in pyelonephritis were associated with the change of the L-forms into bacterial ones, and that the persistence of the L-forms in the kidney tissue promoted the maintenance of the chronic process.