Genetic studies with klebsiella

(1) Numerous attempts were made to detect transformation of Klebsiella strains. In these experiments, DNA was extracted from donor bacteria by a variety of techniques, and the resulting preparations were applied to genetically marked recipient bacteria. In some cases the recipient cells had previously been treated with agents which were known to affect the integrity of the cell wall, and in other cases the recipient cells were taken from ordinary broth cultures. No evidence of transformation was obtained in any experiment; possible reasons for this finding are discussed.(2) Attempts were made to demonstrate lysogeny in Klebsiella aerogenes strain A3> but no positive results were obtained. Similarly, none of the other Klebsiella strains tested were found to release phages active on K. aerogenes strain A3.(3) Several bacteriophages were isolated from natural sources by the enrichment culture technique, with K. aerogenes strain A3 as host. Five of the phages were found to produce turbid zones of partial clearing when plated on strain A3, but none were found to be capable of lysogenising cells of strain A3(4) Attempts were made to transduce K. aerogenes strain A3 and genetically marked derivatives of this strain with a number of virulent phages. Several different methods of minimising superinfection by free phage were used, but no transductants were recovered in any experiment.(5) Auxotrophic mutants of lysogenic and sensitive indicator Klebsiella strains were prepared, and used in preliminary mixed cultivation experiments. The results suggested that genetic exchange had occurred in tests involving K. aerogenes strain W52(6) K. aerogenes strain W52 was found to be lysogenic, but did not appear to be ultraviolet inducible. However, cultures of strain W52 which had been treated with the potent mutagen N-methyl-N'-nitro-N-nitrosoguanidine were found to lyse and release large numbers of plaque-forming particles. This result suggests that NTG is an effective inducing agent, but the possibility that phage lysates obtained by NTGinduction would be unsuitable for use in transduction experiments is discussed.(7) The phage released by strain W52 was propagated in cells of a sensitive strain and designated PW52. Preliminary spot tests suggested that phage PW52 was capable of mediating transduction, with the important limitation that transductants were only recovered if the recipient bacteria were immune to lytic infection by the phage.(8) The characteristics of the system of genetic exchange which appeared to involve phage PW52 were studied, and although absolute confirmation was not obtained the results strongly suggested that phage PW52 was in fact capable _7 of mediating transduction at frequencies varying from 5 x 10 —9 to 3 x 10 per phage particle adsorbed. No evidence was found to suggest that any of the other known mechanisms of genetic exchange were involved in the system which was being studied.(9) It was found that non-mucoid mutants of K. aerogenes strains W52 and W70 could not be transduced with respect to any of the markers tested, and a possible reason for this finding is suggested.(10) Experiments were carried out to test for joint transduction of streptomycin resistance and maltose utilisation markers, but no positive results were obtained.(11) In further preliminary attempts to detect linkage of genetic markers, it was found that certain ade markers appeared to be cotransducible with a strr marker, but later experiments suggested that this result could be explained without postula¬ ting a specific joint transduction of the markers concerned.(12) Some 22 auxotrophic mutants of K. aerogenes strains W52 and W70 were isolated following treatment with N-methyl-N1- nitro-N-nitrosoguanidine, and used in a large series of trans¬ duction tests. The results of these experiments suggested several linkage relationships, which are discussed in some detail. One of the possible linkage relationships was partially confirmed by the donor phenotype selection technique, and several tests which might be expected to provide further confirmation are outlined.(13) Mixed cultivation experiments with auxotrophic derivatives of K. aerogenes strain A3 and K. pneumoniae strain 1.9 failed to yield any evidence of genetic recombination(14) Transfer of the F-lac episome from an E. coli K12 strain to a lac mutant of K. aerogenes strain A3 was observed, but the F-lac+ recombinants were not found capable of trans¬ ferring the episome to an F~lac~ strain of E. coli.(15) Attempts to detect transfer of the F factor from E. coli K12 to Klebsiella strains, or of chromosomal material from an Hfr strain of E. coli K12 to Klebsiella strains, were unsuccessful.(16) Attempts were made to demonstrate elimination of the genetic determinant of bacteriocinogeny from K. pneumoniae strain 1.2 by treatment with acridine dyes, but the results were negative. Experiments designed to detect transfer of the bacteriocin determinant from strain 1.2 to other Klebsiella strains failed to yield any evidence that the determinant was transmissible from cell to cell.(17) It was found that infectious drug resistance (R) factors could be transferred between Klebsiella strains, but transfer was always found to occur at low frequency. The mechanism of the genetic exchange was not characterised fully, but appeared to be a typical example of bacterial conjugation.(18) Experiments involving transfer of an R factor from a capsulate donor strain to capsulate and non-capsulate recipient strains were carried out. No evidence was found to suggest that the non-capsulate strains were capable of acquiring the R factor more efficiently than the capsulate strain.(19) The behaviour of Klebsiella strains in highfrequency resistance transfer systems was studied, and it was concluded that the strains used were not capable of acting as efficient recipients of R factors, and were probably poor donors as well.(20) An R factor which was no longer sensitive to a repressor of its conjugation functions was transferred to Klebsiella strains, and the R+ strains so obtained were used in attempts to detect transfer of chromosomal genetic material to an R~ Klebsiella strain. The R~ strain which was used had been found to act as a reasonably efficient recipient of the derepressed R factor alone, but no chromosomal recombinants were recovered.(21) Derivatives of K. aerogenes strain A3 carrying a derepressed R factor were found to be insensitive to the malespecific bacteriophage MS-2, and appeared to be unable to adsorb significant numbers of MS-2 particles. In control experiments, an E. coli K12 strain carrying the same factor was found to be sensitive to phage MS-2 and capable of removing some 75$ of the MS-2 particles from a culture supernatant

The effect of lipoprotein-associated phospholipase A2 deficiency on pulmonary allergic responses in Aspergillus fumigatus sensitized mice.

BackgroundLipoprotein-associated phospholipase A2 (Lp-PLA2)/platelet-activating factor acetylhydrolase (PAF-AH) has been implicated in the pathogenesis of cardiovascular disease. A therapeutic targeting of this enzyme was challenged by the concern that increased circulating platelet activating factor (PAF) may predispose to or increase the severity of the allergic airway response. The aim of this study was to investigate whether Lp-PLA2 gene deficiency increases the risk of PAF and IgE-mediated inflammatory responses in vitro and in vivo using mouse models.MethodsLp-PLA2-/- mice were generated and back crossed to the C57BL/6 background. PAF-AH activity was measured using a hydrolysis assay in serum and bronchoalveolar lavage (BAL) samples obtained from mice. Aspergillus fumigatus (Af)-specific serum was prepared for passive allergic sensitization of mice in vivo and mast cells in vitro. β- hexosaminidase release was studied in bone marrow derived mast cells sensitized with Af-specific serum or DNP-IgE and challenged with Af or DNP, respectively. Mice were treated with lipopolysaccharide (LPS) and PAF intratracheally and studied 24 hours later. Mice were sensitized either passively or actively against Af and were studied 48 hours after a single intranasal Af challenge. Airway responsiveness to methacholine, inflammatory cell influx in the lung tissue and BAL, immunoglobulin (ELISA) and cytokine (Luminex) profiles were compared between the wild type (WT) and Lp-PLA2-/- mice.ResultsPAF-AH activity was reduced but not completely abolished in Lp-PLA2-/- serum or by in vitro treatment of serum samples with a high saturating concentration of the selective Lp-PLA2 inhibitor, SB-435495. PAF inhalation significantly enhanced airway inflammation of LPS treated WT and Lp-PLA2-/- mice to a similar extent. Sensitized WT and Lp-PLA2-/- bone-marrow derived mast cells released β-hexosaminidase following stimulation by allergen or IgE crosslinking to equivalent levels. Wild type and Lp-PLA2-/- mice responded to passive or active allergic sensitization by significant IgE production, airway inflammation and hyperresponsiveness after Af challenge. BAL cell influx was not different between these strains while IL-4, IL-5, IL-6 and eotaxin release was attenuated in Lp-PLA2-/- mice. There were no differences in the amount of total IgE levels in the Af sensitized WT and Lp-PLA2-/- mice.ConclusionsWe conclude that Lp-PLA2 deficiency in C57BL/6 mice did not result in a heightened airway inflammation or hyperresponsiveness after PAF/LPS treatment or passive or active allergic sensitization and challenge

Optimization of double pulse pumping for Ni-like Sm x-ray lasers

We report a systematic study of double pulse pumping of the Ni-like Sm x-ray laser at 73 Angstrom, currently the shortest wavelength saturated x-ray laser. It is found that the Sm x-ray laser output can change by orders of magnitude when the intensity ratio of the pumping pulses and their relative delay are varied. Optimum pumping conditions are found and interpreted in terms of a simple model. (C) 1999 American Institute of Physics. [S0021-8979(99)07102-9]

Differential involvement of Na(+),K(+)-ATPase isozymes in preimplantation development of the mouse.

Na(+),K(+)-ATPase plays an essential role in mammalian blastocoel formation (cavitation) by driving trans-epithelial sodium transport. Previously, the alpha1 and beta1 subunit isoforms of this enzyme were identified in preimplantation mouse embryos and were assumed to be responsible for this function. Here we show that mRNAs encoding an additional alpha subunit isoform (alpha3) and the remaining two beta subunit isoforms are also present in preimplantation embryos. Whereas alpha3 mRNA accumulates between the four-cell and the blastocyst stages and thus results from embryonic transcription, the same could not be demonstrated for beta2 and beta3 mRNAs. Immunoblot analyses confirmed that these subunits are present in cavitating embryos. Using confocal immunofluorescence microscopy we found that alpha1 and beta1 subunits are concentrated in the basolateral membranes of the trophectoderm while being equally distributed in plasma membranes of the inner cell mass. In contrast, alpha3, beta2, and beta3 subunits were not detected in plasma membranes. Our current assessment, therefore, is that as many as six isozymes of Na(+),K(+)-ATPase could be involved in preimplantation development although it is primarily the alpha1beta1 isozyme that is responsible for blastocoel formation. Our findings imply that the regulation of sodium transport within the preimplantation mouse embryo is more complex than had been appreciated

Optical Coherence Tomography Findings After Childhood Lensectomy

Purpose: To explore the impact of childhood lensectomy on posterior segment development. / Methods: Cross-sectional observational study at children's eye clinics at a tertiary referral center in London, UK. We included 45 children age 4 to 16 years with healthy eyes and 38 who had undergone lensectomy. We acquired posterior segment optical coherence tomography scans of both eyes. We used parametric and nonparametric tests in SPSS24 for the comparison of parameters between groups and within individuals; a P value less than 0.05 was considered significant. The main outcome measures were foveal pit depth and subfoveal choroidal thickness (CT). Secondary outcomes were inner and outer ring CT and photoreceptor layer parameters, macular and peripapillary retinal nerve fiber layer thickness. / Results: Foveal pit depth and subfoveal CT are significantly reduced in eyes that have undergone lensectomy compared with nonoperated eyes. Inner ring CT and outer ring CT are reduced. Foveal inner retinal layer thickness is increased. Mean inner retinal and outer nuclear layer thickness are not affected. / Conclusions: Childhood lensectomy is associated with a reduction in developmental foveal pit deepening and lack of developmental thickening of the posterior choroid. Mechanical and optical disruption of foveal and subfoveal choroidal development may affect structural foveal development after childhood lensectomy

Supersonic strain front driven by a dense electron-hole plasma

We study coherent strain in (001) Ge generated by an ultrafast laser-initiated high density electron-hole plasma. The resultant coherent pulse is probed by time-resolved x-ray diffraction through changes in the anomalous transmission. The acoustic pulse front is driven by ambipolar diffusion of the electron-hole plasma and propagates into the crystal at supersonic speeds. Simulations of the strain including electron-phonon coupling, modified by carrier diffusion and Auger recombination, are in good agreement with the observed dynamics.Comment: 4 pages, 6 figure