Colicinogeny of O55 EPEC diarrhoeagenic Escherichia coli

Approximately 24% of a sample of pathogenic Escherichia coli strains from different serogroups were found to synthesize colicins. Serogroup O55 had an unusually large proportion of such strains (33%). In a sample of 27 O55 isolates, one synthesized a class A colicin (identified as ColE9), five produced class B colicins (three ColIa, two, unidentifiable), and three a class A and a class B together.     

The Plasmid-Encoded Regulator Activates Factors Conferring Lysozyme Resistance on Enteropathogenic Escherichia coli Strains

We demonstrate that enhanced lysozyme resistance of enteropathogenic Escherichia coli requires the plasmid-encoded regulator, Per, and is mediated by factors outside the locus for enterocyte effacement. EspC, a Per-activated serine protease autotransporter protein, conferred enhanced resistance on nonpathogenic E. coli, and a second Per-regulated, espC-independent lysozyme resistance mechanism was identified.     

Drug Resistance of Enteric Bacteria XIII. Distribution of R Factors in Escherichia coli Strains Isolated from Livestock

Escherichia coli strains isolated from 151 swine and 108 fowl, which were kept at the Animal Health Center, Maebashi, Japan, were surveyed for drug resistance and distribution of R factors. All of the swine and 38% of the fowl excreted E. coli strains resistant to tetracycline, chloramphenicol, streptomycin, and sulfanilamide, or certain combinations thereof. Among 278 resistant cultures isolated from swine, 13% were found to be resistant to one antibiotic, whereas 87% were resistant to more than one antibiotic. Among these resistant strains, 40% carried R factors which were transferable by the usual conjugal process. The resistance patterns of these R factors included 36% which were singly resistant and 64% which were multiply resistant. Among 54 resistant cultures isolated from fowl, 24% were singly resistant and 76% were multiply resistant. Of the resistant strains from fowl, 22% carried R factors. The resistance patterns of R factors included 50% of the singly resistant type and 50% which were multiply resistant. In spite of feeding with dairy products containing only tetracycline, a high incidence of multiple resistance was observed in the E. coli strains and the R factors isolated from these animals.     

Resistance to Arsenic Compounds Conferred by a Plasmid Transmissible Between Strains of Escherichia coli

A plasmid, R773, which confers resistance to arsenic compounds, is transmissible between strains of Escherichia coli. It is a member of compatibility group F(1).     

Biotic and Abiotic Factors Affecting Plasmid Transfer in Escherichia coli Strains

[This corrects the article on p. 393 in vol. 58.].     

Loss of Lysis Inhibition in Filamentous Escherichia coli Infected with Wild-Type Bacteriophage T4

The plaque enlargement of wild-type T4 bacteriophage observed when assayed in the presence of low concentrations of mitomycin C or after exposure to very low doses of ultraviolet light was studied by using solid as well as liquid culture media. It was found that the filamentous cell formed by the treatment with the agents is responsible for the phenomenon. The filamentous cell was also shown to be characterized not only by the loss of capacity of lysis inhibition but also by a shortening of the latent period. No difference in cellular rigidity could be seen between the filamentous cell and normal cell as far as the analysis from the outside of the cell was concerned, whereas the former cell was shown to be more readily susceptible to phage-induced lysozyme from the inside of the cell. A possible change in the membrane of the filamentous cell and a possible mechanism for lysis inhibition are discussed.     

Combinatorial Strategies for Improving Multiple-Stress Resistance in Industrially Relevant Escherichia coli Strains

High-cell-density fermentation for industrial production of chemicals can impose numerous stresses on cells due to high substrate, product, and by-product concentrations; high osmolarity; reactive oxygen species; and elevated temperatures. There is a need to develop platform strains of industrial microorganisms that are more tolerant toward these typical processing conditions. In this study, the growth of six industrially relevant strains of Escherichia coli was characterized under eight stress conditions representative of fed-batch fermentation, and strains W and BL21(DE3) were selected as platforms for transposon (Tn) mutagenesis due to favorable resistance characteristics. Selection experiments, followed by either targeted or genome-wide next-generation-sequencing-based Tn insertion site determination, were performed to identify mutants with improved growth properties under a subset of three stress conditions and two combinations of individual stresses. A subset of the identified loss-of-function mutants were selected for a combinatorial approach, where strains with combinations of two and three gene deletions were systematically constructed and tested for single and multistress resistance. These approaches allowed identification of (i) strain-background-specific stress resistance phenotypes, (ii) novel gene deletion mutants in E. coli that confer single and multistress resistance in a strain-background-dependent manner, and (iii) synergistic effects of multiple gene deletions that confer improved resistance over single deletions. The results of this study underscore the suboptimality and strain-specific variability of the genetic network regulating growth under stressful conditions and suggest that further exploration of the combinatorial gene deletion space in multiple strain backgrounds is needed for optimizing strains for microbial bioprocessing applications.     

Homolactate Fermentation by Metabolically Engineered Escherichia coli Strains

We report the homofermentative production of lactate in Escherichia coli strains containing mutations in the aceEF, pfl, poxB, and pps genes, which encode the pyruvate dehydrogenase complex, pyruvate formate lyase, pyruvate oxidase, and phosphoenolpyruvate synthase, respectively. The process uses a defined medium and two distinct fermentation phases: aerobic growth to an optical density of about 30, followed by nongrowth, anaerobic production. Strain YYC202 (aceEF pfl poxB pps) generated 90 g/liter lactate in 16 h during the anaerobic phase (with a yield of 0.95 g/g and a productivity of 5.6 g/liter · h). Ca(OH)₂ was found to be superior to NaOH for pH control, and interestingly, significant succinate also accumulated (over 7 g/liter) despite the use of N₂ for maintaining anaerobic conditions. Strain ALS961 (YYC202 ppc) prevented succinate accumulation, but growth was very poor. Strain ALS974 (YYC202 frdABCD) reduced succinate formation by 70% to less than 3 g/liter. ¹³C nuclear magnetic resonance analysis using uniformly labeled acetate demonstrated that succinate formation by ALS974 was biochemically derived from acetate in the medium. The absence of uniformly labeled succinate, however, demonstrated that glyoxylate did not reenter the tricarboxylic acid cycle via oxaloacetate. By minimizing the residual acetate at the time that the production phase commenced, the process with ALS974 achieved 138 g/liter lactate (1.55 M, 97% of the carbon products), with a yield of 0.99 g/g and a productivity of 6.3 g/liter · h during the anaerobic phase.     

Drug Resistance and R Plasmids in Escherichia coli Strains Isolated from Imported Pet Birds

Drug resistance in Escherichia coli strains isolated from pet birds (mynahs, macaws, finches, common bengals, parrots, and flamingos) imported into Japan from 10 foreign countries in 1977 and 1978 was investigated. Of the 309 strains isolated from 127 pet birds in the Animal Quarantine Service, 232 (75.1%) were drug resistant. Furthermore, strains resistant to oxytetracycline hydrochloride, dihydrostreptomycin, and sulfadimethoxine were relatively common. Resistance patterns varied from single to sextuple resistance, and 148 (63.8%) of the resistant strains had conjugative R plasmids. These results suggest that the high incidence of drug resistance and R plasmids in E. coli strains isolated from these pet birds may be a reflection of the prophylactic use of antibiotics for the prevention of diseases which increasingly occur with importation of the birds. Furthermore, the results suggest that the birds may be potential reservoirs of drug-resistant E. coli for families who raise and have intimate contact with such birds.     

Chromosomal Integration of F′ Factors in Recombination-Deficient Hfr Strains of Escherichia coli

In matings between F′ donors and recombination-deficient Hfr recipients, we isolated progeny which transferred both episomal markers and Hfr markers early and with high frequency. A number of these progeny had two integrated sex factors. Investigation of these double Hfr strains showed that the F′ nearly always integrated in a homologous region of the chromosome. In any particular mating system integration was specific as to location and direction of chromosome transfer.     

Genetic Stability of Various Resistance Factors in Escherichia coli and Salmonella typhimurium

Genetic stability of R factors was studied in Salmonella typhimurium LT-2 and Escherichia coli K-12. It was found that fi(+) R [or R(f)] factors were unstable in LT-2, losing their drug-resistance markers at high frequencies, and were stable in K-12; fi(-) R [or R(i)] factors were stable in both hosts. Both fi(+) and fi(-) R factors were genetically stable also in recombination-deficient mutants of K-12. An fi(+) R factor, which was unstable in S. typhimurium LT-2 wild type, was relatively stable in a recombination-deficient mutant of LT-2. In the spontaneous loss of the drug-resistance markers of fi(+) R factors in LT-2, the markers for sulfanilamide, streptomycin, and chloramphenicol resistance were lost together at high frequencies and the tetracycline marker was retained stably. The remaining drug-resistance markers of the spontaneous segregants of LT-2 were transmissible to K-12 by mixed cultivation, indicating that they were still in the form of R factors.     

Cytochemical Localization of Certain Phosphatases in Escherichia coli

Cytochemical studies of Escherichia coli at the light and electron microscopic levels have revealed alkaline phosphatase, hexose monophosphatase, and cyclic phosphodiesterase reaction products in the periplasmic space and at the cell surface. In preparations for both light and electron microscopy, reaction product filled polar caplike enlargements of the periplasmic space, such as those described in plasmolyzed cells, indicating significant terminal concentrations of these enzymes; dense substance was often seen within these polar caps in morphological specimens. Staining of the bacterial surface was commonly encountered, but could represent artifactual accumulation of precipitate along the cell wall. Alkaline phosphatase was demonstrated with several substrates (ethanolamine phosphate, glycerophosphate, p-nitrophenylphosphate, and glucose-6-phosphate) over a wide pH range in a bacterial strain (C-90) known to be constitutive for this enzyme, whereas strains deficient in this enzyme (U-7, repressed K-37), showed no activity with these substrates. Hexose monophosphatase and cyclic phosphodiesterase activities were characterized by reaction-product deposition with specific substrates at acid or neutral, but not at alkaline, pH in strains of E. coli lacking alkaline phosphatase (U-7 and repressed K-37). Fixation in Formalin or the use of calcium as a capture reagent seemed to interfere with periplasmic staining in cells prepared for electron microscopy. Formalin fixation had little effect on biochemical assays of the phosphatase activity of intact cells in suspension, but partially reduced the activity evident in sonically treated extracts or in suspensions of dispersed cryostat sections. Glutaraldehyde treatment impaired enzyme activity more drastically.     

峨眉山藏猕猴粪源大肠杆菌耐药性的调查研究

目的了解峨眉山藏猕猴猴群大肠杆菌耐药性情况,为疫病防治和耐药性在野生猕猴群体中的扩散提供科学依据。方法采集峨眉山藏猕猴粪便105份,进行大肠杆菌的分离鉴定,并对分离杆菌进行致病性试验以及按照CLSI推荐的K-B药敏纸片法进行19种抗生素的耐药性检测。结果从105份样品中分离到105株大肠杆菌,其中6株致病性较强,23株有致病性,76株无致病性。各景区所分离的大肠杆菌对痢特灵(FUZ)耐药率较高,一线天达54.29%,钻天坡达39.47%,雷洞坪达46.87%。105株不同区域来源的大肠杆菌耐受的药物种数不同,一线天景区无12耐以上的菌株,而以3耐最多,为11.43%。钻天坡景区无7耐以上的菌株,而以1耐最多,为21.05%。雷洞坪景区无8耐以上的菌株,而以1耐最多,为18.75%。对各猴区分离的29株致病性大肠杆菌对19种抗生素共产生了7种耐药谱。总体来看,优势耐药谱不明显,且不同个体致病性大肠杆菌分离株的耐药谱存在差异。结论应定期追踪峨眉山藏猕猴大肠杆菌耐药情况,对野生藏猕猴疫病的防控具有比较重要的意义。     

Glycolate Uptake by Mutant Strains of Escherichia coli K-12

Mutant strains of Escherichia coli K-12 were shown to be impaired in their ability to assimilate glycolate-2-(14)C. One strain (Glc-103) has lost the ability to oxidize glycolate; another strain (Glc-102) was relatively impermeable to the compound. A third strain (Glc-104) had undergone a similar loss in permeability, and, in addition, was deranged in the synthesis of either glyoxylate reductase or malate synthase G.     

Escherichia coli with Resistance Factors in Vegetarians, Babies, and Nonvegetarians

The prevalence of Escherichia coli carrying resistance factors (R factors) was examined in meat-consuming individuals and in those not consuming meat (vegetarians and babies below the age of 6 months). Assuming that the transport of resistant E. coli from animals through meat and meat products to the human consumer is most important, with regard to the incidence of resistant E. coli in man, we expected a significant difference in the proportions of people with resistant E. coli between the two groups. However, the percentage with resistant E. coli was larger in the group of vegetarians and babies than in the group of meat-eating individuals.     

Dissemination of Cephalosporin Resistance Genes between Escherichia coli Strains from Farm Animals and Humans by Specific Plasmid Lineages

Third-generation cephalosporins are a class of β-lactam antibiotics that are often used for the treatment of human infections caused by Gram-negative bacteria, especially Escherichia coli. Worryingly, the incidence of human infections caused by third-generation cephalosporin-resistant E. coli is increasing worldwide. Recent studies have suggested that these E. coli strains, and their antibiotic resistance genes, can spread from food-producing animals, via the food-chain, to humans. However, these studies used traditional typing methods, which may not have provided sufficient resolution to reliably assess the relatedness of these strains. We therefore used whole-genome sequencing (WGS) to study the relatedness of cephalosporin-resistant E. coli from humans, chicken meat, poultry and pigs. One strain collection included pairs of human and poultry-associated strains that had previously been considered to be identical based on Multi-Locus Sequence Typing, plasmid typing and antibiotic resistance gene sequencing. The second collection included isolates from farmers and their pigs. WGS analysis revealed considerable heterogeneity between human and poultry-associated isolates. The most closely related pairs of strains from both sources carried 1263 Single-Nucleotide Polymorphisms (SNPs) per Mbp core genome. In contrast, epidemiologically linked strains from humans and pigs differed by only 1.8 SNPs per Mbp core genome. WGS-based plasmid reconstructions revealed three distinct plasmid lineages (IncI1- and IncK-type) that carried cephalosporin resistance genes of the Extended-Spectrum Beta-Lactamase (ESBL)- and AmpC-types. The plasmid backbones within each lineage were virtually identical and were shared by genetically unrelated human and animal isolates. Plasmid reconstructions from short-read sequencing data were validated by long-read DNA sequencing for two strains. Our findings failed to demonstrate evidence for recent clonal transmission of cephalosporin-resistant E. coli strains from poultry to humans, as has been suggested based on traditional, low-resolution typing methods. Instead, our data suggest that cephalosporin resistance genes are mainly disseminated in animals and humans via distinct plasmids.     

New Flagellin-Specifying Genes in Some Escherichia coli Strains

Data for further development of the flagellar antigen genetics of the species Escherichia coli are reported. Two new flagellin genes named fllA and flmA were found in E. coli 781-55, E2987-73, and E223-69, the test strains for E. coli flagellar antigens H44, H55, and H54, respectively (collection of the International Escherichia and Klebsiella Centre of the World Health Organization, Copenhagen, Denmark). Two alleles of fllA were identified that encode flagellar antigens H44 (fllA₄₄) and H55 (fllA₅₅), and the only flmA allele found (flmA₅₄) encodes antigen H54. The sites of their integration in the E. coli K-12 chromosome after P1-mediated transduction were approximately determined and found to be separate from each other and from the known regions of flagellar genes of E. coli and salmonellae. The region of flm₅₄ was found to repress the expression of some alleles of the flagellin gene fliC. In addition, cryptic genes encoding antigens H4 and H38 were found in phenotypically monophasic test strains 781-55 and E2987-73, respectively.     

Elimination of Sex Factors in Escherichia coli by Urea

Eliminatory action of urea on the sex factor (F) in Escherichia coli K-12 strains is reported. Growth of E. coli harboring F or F'8 (F-gal) factors in Penassay Broth containing urea led to the loss of these genetic elements and yielded F(-) cells. Appearance of F(-) cells among survivors was already observed when the culture was in the very early stage of exponential phase. However, frequencies of F(-) cells formed did not increase much as a function of the incubation time. Unusual F(+) or F'8 cells which retained the ability of genetic transfer but showed resistance to M12 phage were also isolated. Addition of sucrose to broth with urea led to the favorable growth of cells in the culture and the increase, if little, of elimination frequencies of F factors by urea. These findings, coupled with other observations, suggest that urea has two separate actions in enhancing the frequency of F(-) bacteria, namely, (i) to inactivate F by direct action, such as mutation, and (ii) to select the F(-) variants by differentially inhibiting the growth of F(+).     

Emergence of Variability in Isogenic Escherichia coli Populations Infected by a Filamentous Virus

The spread of epidemics not only depends on the average number of parasites produced per host, but also on the existence of highly infectious individuals. It is widely accepted that infectiousness depends on genetic and environmental determinants. However, even in clonal populations of host and viruses growing in homogeneous conditions, high variability can exist. Here we show that Escherichia coli cells commonly display high differentials in viral burst size, and address the kinetics of emergence of such variability with the non-lytic filamentous virus M13. By single-cell imaging of a virally-encoded fluorescent reporter, we monitor the viral charge distribution in infected bacterial populations at different time following infection. A mathematical model assuming autocatalytic virus replication and inheritance of bacterial growth rates quantitatively reproduces the experimental distributions, demonstrating that deterministic amplification of small host inhomogeneities is a mechanism sufficient to explain large and highly skewed distributions. This mechanism of amplification is general and may occur whenever a parasite has an initial phase of exponential growth within its host. Moreover, it naturally reproduces the shift towards higher virulence when the host is experimenting poor conditions, as observed commonly in host-parasite systems.