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Abstract Since Ukraine is a powerful maize exporter in the world, screening genotypes susceptible to genetic transformation among those cult

Abstract Since Ukraine is a powerful maize exporter in the world, screening genotypes susceptible to genetic transformation among those cultivated on its territory and the development of an effective technology for the transformation of maize of Ukrainian breeding are relevant. Precultivated immature maize embryos of nine genotypes (inbred lines and hybrids) were subjected to Agrobacterium-mediated transformation. Three strains of A. tumefaciens and three vector constructs, which contained the selective marker gene of neomycin phosphotransferase II, as well as the reporter genes of green fluorescent protein or β-glucuronidase, were used in the work. Transgenic plants were selected on nutrient media with paromomycin. For six genotypes, shoot regeneration was observed after transformation on selective media. The presence of transgenes in 43 clones obtained from four genotypes using all available vectors was revealed by the PCR method. The transformation frequency varied from 0 to 27% in individual experiments. A strong correlation was found between frequencies of the regeneration and the transformation between genotypes and donor plants of the same genotype. The use of the pCB271 vector, which contained monocot nucleotide sequences of gene expression regulation upstream of the selective marker gene, namely the intron of the maize hsp70 gene, significantly increased the frequency of shoot regeneration after transformation. Analysis of T1 generation plants revealed single-locus integration of transgenes into the plant genome. The proposed protocol of genetic transformation using paromomycin as a selective agent is effective in obtaining transgenic maize lines and hybrids of Ukrainian breeding. Three genotypes (inbred line DK232 and hybrids KP7×PRZh5 and KS277×RS15) were selected as the most susceptible among tested genotypes to Agrobacterium-mediated transformation and promising for further research on the production of transgenic maize plants.

#Zeamays #nptII #S65Tpgfp #uidA #paromomycin #transgenicmaize #generationT1 #Zea #maize #Agrobacterium #UkrainianBreeding #Ukrainiane

Giardiasis – Causes, Symptoms, Diagnosis, Treatment and Ongoing care

Intestinal infection caused by the protozoan parasite Giardia lamblia (1); G. lamblia is also called G. duodenalis and G. intestinalis. Infection results from ingestion of the cysts, which excyst into trophozoites. These colonize the small intestine and cause symptoms. Giardiasis – Causes, Symptoms, Diagnosis, Treatment and Ongoing care

#contaminated water #Crohn #Diarrhea #Lactose intolerance #Paromomycin #protozoan parasite #public swimming pools #small intestine #water purification #G #Intestinal

To Treat or Not To Treat... But How?

In the "To Treat or Not To Treat" series (please look up previous post here), we have come to the "...But How?" episode. Blastocystis may be susceptible to a number of drugs - in vitro. In vitro is not the opposite of in vivo. In vitro just means that the test has been done on an organism that has been isolated from its usual habitat and tested e.g. in a flask, test tube, etc. In the lab, strains can be challenged and manipulated in multiple ways, but there is no guarantee that the outcome of an in vitro susceptibility test is reproducible in vivo, i.e. when the organism is challenged in its natural habitat and under "natural" conditions. Hence, if you test Blastocystis against metronidazole or any other compound (such as iodine) in vitro, and you observe an effect, you cannot rely on being able to reproduce the effect in vivo. This is due to a variety of reasons including pharmaco-kinetics and pharmaco-dynamics, including the ability of the drug to reach the parasite in its ecological niche, impact of the drug on other micro-organisms, drug interactions, strain-dependent differences in susceptibility (including inherent or acquired resistance), etc. We recently described a case in which a woman with irritable bowel syndrome (according to the Rome III criteria) had both Blastocystis subtype 9 (ST9) and Dientamoeba fragilis. In order to try and eradicate the parasites and to see whether any eradication would impact on her clinical situation, she received multiple courses of antibiotic treatment: 1. Metronidazole (750 mg x 3/d for 10 days) 2. Tetracycline (500 mg x 4/d for 10 days) 3. Trimethoprim + Sulfamethoxazole (TMP 800 mg + SXT 160 mg x 2/d for 7 days) 4. Mebendazole + Metronidazole (100 mg x 2 separated by 2 weeks; subsequently metronidazole as in 1.) 5. Paromomycin + Metronidazole (PM 500 mg + MZ 170 mg x 3/d for 10 days) Mebendazole was given to the entire household due to suspicion of pinworm infection running in the family that could be a potential reservoir of D. fragilis (re-)infection. No clinical alleviation was seen throughout this period. PCR-based detection of Blastocystis and D. fragilis was used to evaluate faecal samples 5-10 days post-treatment: Microbiological effect was seen only on D. fragilis which was cleared only after treatment with PM + MZ (5). So, Blastocystis "survived" this series of antimicrobial treatment. In Denmark, no further relevant treatment options are available for general use (actually, even the use of Humatin (PM) needs a special license). None of the patient's family members or pets were found to be colonised by the same strain, probably indicating that there was no "local" reservoir for ST9, and that the repeated finding of ST9 was not due to re-infection. It may be so that Blastocystis requires a certain intestinal bacterial flora to establish. However, we expect that substantial perturbations in the intestinal flora must have taken place during the patient's various treatments, and therefore Blastocystis must be able to quickly overcome and adapt to such perturbations. It may add to the conundrum that in this case the woman harboured ST9, which is only very rarely seen in humans, and we might therefore deduce that its presence would be more volatile. No animal/environmental reservoir has yet been identified for ST9. There is no doubt that microbiomic profiling of the intestinal flora would be of great benefit in a case like this. If data could be achieved on the impact of these drugs on the relative bacterial structure and function by metagenomic approaches, then this would allow us to explore the changes in the general flora that Blastocystis is capable of withstanding. Certainly, none of these drugs had a measurable in-vivo protistocidal effect on Blastocystis when administered as shown. I re-emphasise that it is far from certain that Blastocystis is capable of inducing disease, directly or indirectly, and hence, we do not know if, and in which situations, we should aim at eradicating it. Suffice it to say, that in our hands and with the compounds that are available for general use in Denmark, it is apparently extremely challenging to eradicate Blastocystis, if at all possible.

Microbe Resilience (Source)

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