Transferable colicinogenic factors as mobilizing agents for extrachromosomal streptomycin resistance

Fredericq, P; Krčméry, V.; Kettner, M.

doi:10.1002/jobm.3630110103

Cited by 12 publications

(1 citation statement)

References 11 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…However, the low-copy, broad-host-range replicon RSF1010, originally isolated from E. coli [38], was not known to be associated with heterogeneity until now. And replication systems with an even lower average PCN (RK2, pBBR1, pSC101) did not show any bimodality under the same conditions in this study.…”

Section: Discussionmentioning

confidence: 99%

Copy number variability of expression plasmids determined by cell sorting and Droplet Digital PCR

et al. 2016

View full text Add to dashboard Cite

BackgroundPlasmids are widely used for molecular cloning or production of proteins in laboratory and industrial settings. Constant modification has brought forth countless plasmid vectors whose characteristics in terms of average plasmid copy number (PCN) and stability are rarely known. The crucial factor determining the PCN is the replication system; most replication systems in use today belong to a small number of different classes and are available through repositories like the Standard European Vector Architecture (SEVA).ResultsIn this study, the PCN was determined in a set of seven SEVA-based expression plasmids only differing in the replication system. The average PCN for all constructs was determined by Droplet Digital PCR and ranged between 2 and 40 per chromosome in the host organism Escherichia coli. Furthermore, a plasmid-encoded EGFP reporter protein served as a means to assess variability in reporter gene expression on the single cell level. Only cells with one type of plasmid (RSF1010 replication system) showed a high degree of heterogeneity with a clear bimodal distribution of EGFP intensity while the others showed a normal distribution. The heterogeneous RSF1010-carrying cell population and one normally distributed population (ColE1 replication system) were further analyzed by sorting cells of sub-populations selected according to EGFP intensity. For both plasmids, low and highly fluorescent sub-populations showed a remarkable difference in PCN, ranging from 9.2 to 123.4 for ColE1 and from 0.5 to 11.8 for RSF1010, respectively.ConclusionsThe average PCN determined here for a set of standardized plasmids was generally at the lower end of previously reported ranges and not related to the degree of heterogeneity. Further characterization of a heterogeneous and a homogeneous population demonstrated considerable differences in the PCN of sub-populations. We therefore present direct molecular evidence that the average PCN does not represent the true number of plasmid molecules in individual cells.Electronic supplementary materialThe online version of this article (doi:10.1186/s12934-016-0610-8) contains supplementary material, which is available to authorized users.

show abstract

Section: Discussionmentioning

confidence: 99%

Copy number variability of expression plasmids determined by cell sorting and Droplet Digital PCR

et al. 2016

View full text Add to dashboard Cite

show abstract

Transferable resistance to tetracycline in Salmonella anatum

Štepánková¹,

Krčméry²,

Janousková³

1971

J of Basic Microbiology

View full text Add to dashboard Cite

During obligatory bacteriological examinations of persons occupied in food industry or distribution, we isolated from the stool specimens of 3 persons of a meat factory strains of Salmonella anatum resistant to antibiotics.Two atrains, S. anatum 5915 and 8. anatum 6100, respectively, were found to be resistant to tetracyclines (T) only, while the third strain showed an unusual type of resistance to colistine (colimycin).Our Antibiotic Reference Laboratory has just started a thorough examination of resistant Salmonella strains in Czechoslovakia for the presence of R-factors.Both T-resistent strains were therefore tested, in mixed cultures, with Escherichia coli K 12 185 resistant to nalidixic acid (Nx), for t&e transfer of their T-monoresistance. The procedure we are using for the detection of R-factors and for following up their further transfers, was already described (KRCMERY and J A N O U~K O V~ 1969, FRI~DERICQ et al. 1971). Briefly, 2 ml of 6 hrs. shaken cultures of the donor strains in nutrient broth (DIFCO) were mixed with equal volumes of 6 hrs. broth cultures of E. coli K 12 185 Nx, which is susceptible to T, but has a chromosomally located high-level resistance to Nx. The strains of S. anatum tested were all highly sensitive to Nx as seen in control plating on MACCONKEY Agar (DIFCO) containing 0.5 to 10.0 pglml of Nx. After overnight incubation, the mixture was diluted to lo-' in saline, and 0.01 ml of each dilution was plated on a segment of MACCONKEY Agar containing 20 pg/ml of T and the same amount of Nx. The plates were incubated 20 hrs. a t 37 "C.The growth with a considerable frequency on MACCONKEY Agar with T and Nx of E. coli K 12 185 Nx after the mixed incubation with 8. anaturn strain 5915suggests the transferable nature of the T-determinant in that Salmonella strain.The T-determinant of S.anatum 6100 was not transferable in the experimental system used, which, however, does not exclude its transferability t o another recipient strain. The frequencies of transfer are given as the fraction of resistant colonies of the recipient strain out of the total number of recipient cells, which was about 2.8 x lo8 cells/ml. I n the above instance, about los cells/ml of the recipient strain, present in the mixture with S. anatum 5915, received the Tdeterminant. The level of acquired T-resistance in the colonies of strain E. coli K 12 185 Nx on MACCONKEY Agar was between 50 and 75 pg/ml of T.The transferability of this T-determinant was reconfirmed by its further transfer from E. coli K 12 185 Nx T to S. typhimurium SH, susceptible to T

show abstract