Hybridization alters spontaneous mutation rates in a parent-of-origin-dependent fashion in Arabidopsis

Over 70 years ago, increased spontaneous mutation rates were observed in Drosophila spp. hybrids, but the genetic basis of this phenomenon is not well understood. The model plant Arabidopsis (Arabidopsis thaliana) offers unique opportunities to study the types of mutations induced upon hybridization and the frequency of their occurrence. Understanding the mutational effects of hybridization is important, as many crop plants are grown as hybrids. Besides, hybridization is important for speciation and its effects on genome integrity could be critical, as chromosomal rearrangements can lead to reproductive isolation. We examined the rates of hybridization-induced point and frameshift mutations as well as homologous recombination events in intraspecific Arabidopsis hybrids using a set of transgenic mutation detector lines that carry mutated or truncated versions of a reporter gene. We found that hybridization alters the frequency of different kinds of mutations. In general, Columbia (Col) × Cape Verde Islands and Col × C24 hybrid progeny had decreased T→G and T→A transversion rates but an increased C→T transition rate. Significant changes in frameshift mutation rates were also observed in some hybrids. In Col × C24 hybrids, there is a trend for increased homologous recombination rates, except for the hybrids from one line, while in Col × Cape Verde Islands hybrids, this rate is decreased. The overall genetic distance of the parents had no influence on mutation rates in the progeny, as closely related accessions on occasion displayed higher mutation rates than accessions that are separated farther apart. However, reciprocal hybrids had significantly different mutation rates, suggesting parent-of-origin-dependent effects on the mutation frequency

Bioactivity Studies of β-Lactam Derived Polycyclic Fused Pyrroli-Dine/Pyrrolizidine Derivatives in Dentistry: In Vitro, In Vivo and In Silico Studies.

The antibacterial activity of β-lactam derived polycyclic fused pyrrolidine/pyrrolizidine derivatives synthesized by 1, 3-dipolar cycloaddition reaction was evaluated against microbes involved in dental infection. Fifteen compounds were screened; among them compound 3 showed efficient antibacterial activity in an ex vivo dentinal tubule model and in vivo mice infectious model. In silico docking studies showed greater affinity to penicillin binding protein. Cell damage was observed under Scanning Electron Microscopy (SEM) which was further proved by Confocal Laser Scanning Microscope (CLSM) and quantified using Flow Cytometry by PI up-take. Compound 3 treated E. faecalis showed ROS generation and loss of membrane integrity was quantified by flow cytometry. Compound 3 was also found to be active against resistant E. faecalis strains isolated from failed root canal treatment cases. Further, compound 3 was found to be hemocompatible, not cytotoxic to normal mammalian NIH 3T3 cells and non mutagenic. It was concluded that β-lactam compound 3 exhibited promising antibacterial activity against E. faecalis involved in root canal infections and the mechanism of action was deciphered. The results of this research can be further implicated in the development of potent antibacterial medicaments with applications in dentistry

Time kill assay for <i>E</i>. <i>faecalis</i>.

<p>Time kill assay confirmed growth inhibitory effect of <i>β-</i>lactam compounds (3, 7 and 6a) against <i>E</i>. <i>faecalis</i>. Samples were collected at the indicated times and were evaluated for CFUs. Time kill kinetics of <i>E</i>. <i>faecalis</i> by compound 3 (25 μg/ml) represent dead cells within 1 h. Note: Ut- untreated, Amp- Ampicillin.</p

Mammalian cell cytotoxicity.

<p>MTT based cytotoxicity assay on NIH 3T3. The cells were treated with various concentrations of <i>β-</i>lactam compounds and incubated for 24 h. After incubation, the percentage of cell viability was calculated. Standard deviations from three observations were plotted. Note: Amp- Ampicillin.</p

Synthesis of <i>β-</i>lactam Baylis-Hillman adducts.

<p>Synthesis of <i>β-</i>lactam Baylis-Hillman adducts.</p

<i>In vivo</i> anti bacterial efficacy of <i>β-</i>lactam in <i>E</i>. <i>faecalis</i> infected mice.

<p>Five mice in each group received three subcutaneous doses of <i>β-</i>lactams (25 and 50 mg/kg) and ampicillin (25 mg/kg) after 4 h of intravenous inoculation of <i>E</i>. <i>faecalis</i>. 120 h after infection and daily treatment with dose indicated, the kidney were harvested and CFU were measured in kidney. Non-parametric test was done for comparisons of significance for test versus control (<i>p</i>< 0.05).</p><p><i>In vivo</i> anti bacterial efficacy of <i>β-</i>lactam in <i>E</i>. <i>faecalis</i> infected mice.</p

Minimum Inhibitory Concentration (μg/ml) of <i>β</i>-lactams against pathogens in root canal infection.

<p>-, no activity, Amp- Ampicillin, RS (1–5)—Resistant strains isolated from Root Canal Treatment failure cases.</p><p>Minimum Inhibitory Concentration (μg/ml) of <i>β</i>-lactams against pathogens in root canal infection.</p

Synthesis of <i>β-</i>lactam substituted polycyclic fused pyrrolidine/pyrrolizidine cycloadducts.

<p>Synthesis of <i>β-</i>lactam substituted polycyclic fused pyrrolidine/pyrrolizidine cycloadducts.</p

Biofilm assay.

<p>Qualitative SEM study to show inhibition of biofilm formation. Images show reduction of biofilm after 24 h treatment with agents A) Large number of viable adherent bacterial cells on the matrix (control without treatment) B) bacterial reduction after treatment with <i>β-</i>lactam compound 3 (25 μg/ml) C) bacterial reductions after treatment with ampicillin. Notes: scale bar: 2 nm. D) Quantitative spectrophotometric measurement to show inhibition of biofilm formation. Graph indicates percentage of biofilm reduction after 24 h treatment with respective agent. Nonparametric test was done for comparisons of significance for test versus control (<i>p</i>< 0.05). Note: Amp- Ampicillin.</p