Using the structural diversity of siderophore biosynthesis genes for intra-and interspecific differentiation of pathogenic yersinia

2021 ◽  
pp. 50-56
Author(s):  
Daria Aleksandrovna Kuznetsova ◽  
Aleksey Leonidovich Trukhachev ◽  
Violetta Aleksandrovna Rykova ◽  
Olga Nikolaevna Podladchikova
Keyword(s):  
Tandem Repeats ◽  
Variable Structure ◽  
Structural Diversity ◽  
Variable Number ◽  
Differential Diagnostics ◽  
Pcr Analysis ◽  
Unknown Variable ◽  
Siderophore Biosynthesis ◽  
Pathogenic Yersinia

The paper analyzes the siderophore biosynthesis genes that are located in the ysu and ynp loci of only Y. pestis and Y. pseudotuberculosis, have variable structure between different strains of both species and contain previously unknown variable number tandem repeats (VNTR). The purpose of the study was to assess the possibility of using these VNTR as genetic markers for intra-and interspecific differentiation of pathogenic Yersinia. Based on the novel VNTR-markers, three pairs of primers (ysu-interF/R, ilp1F/R и ilp2F/R) were designed and used for the in silico and in vitro PCR analysis of various Y.pestis and Y. pseudotuberculosis strains. All studied Y. pestis strains of the main subspecies (ssp pestis), unlike the strains of non-main subspecies and Y. pseudotuberculosis, did not give amplicon with ilp1F/R primers, since the area between them contains an IS100 element. To identify the strains of the main subspecies, the fourth pair of primers ilp1F-is100R was designed, allowing the most dangerous ssp pestis strains to be distinguished from the not dangerous non-main ssp strains. Y. pseudotuberculosis strains were characterized by a significant variety of amplicons with three pairs of primers, and which made it possible to carry out intraspecies strain genotyping. At the same time, for those strains whose serotype is known, the correlation between the serogroup and the genotype of the strains was observed. Analysis of the 1 serotype strains representing most sequenced Y. pseudotuberculosis strains allowed us to separate two gene groups differing from the rest of 1 serotype gene groups. The first one included the serotype 1a strains isolated from people in Europe, which are known to have the greatest pathogenetic potential. The other one was formed by serotype 1b strains isolated from people in Siberia and Primorye, which are characterized by the high epidemic potential. Thus, four pairs of primers designed in this study can be used to develop additional tests for the identification and differential diagnostics of the most dangerous Y. pestis ssp pestis and Y. pseudotuberculosis serotype 1a and 1b strains.

Introduction of Infected Animals to Herds Is an Important Route for the Spread of Yersinia enterocolitica Infection between Pig Farms

2014 ◽  
Vol 77 (1) ◽  
pp. 116-121 ◽  
Author(s):  
S. VIRTANEN ◽  
S. NIKUNEN ◽  
H. KORKEALA
Keyword(s):  
Yersinia Enterocolitica ◽  
Tandem Repeats ◽  
Variable Number ◽  
Multiple Locus ◽  
Pig Farms ◽  
Sources Of Infection ◽  
Enterocolitica Infection ◽  
Variable Number Tandem Repeats ◽  
Pathogenic Yersinia

Altogether, 369 pathogenic Yersinia enterocolitica isolates from 1,118 fecal samples collected from 22 pig farms of different production types were characterized by biotyping, serotyping, and genotyping using multiple-locus variable-number tandem repeats analysis. We investigated the distribution of the different genotypes at the farm level and their association with different farm conditions. Pigs were found to carry and transmit Y. enterocolitica between farms, because the same genotypes were found on farms that had previously transported the pigs between them. The purchase of new animals for the farms associated significantly with the number of different multiple-locus variable-number tandem repeats analysis types of Y. enterocolitica found within a farm. Some genotypes seemed to persist on farms for years. The results of this study show that pigs purchased from infected herds transmit Y. enterocolitica infection between farms. Certain pig farms may act as long-term sources of infection.

Suitability of loci for multiple-locus variable-number of tandem-repeats analysis of Cryptosporidium parvum for inter-laboratory surveillance and outbreak investigations

Parasitology ◽  
2016 ◽  
Vol 144 (1) ◽  
pp. 37-47 ◽  
Author(s):  
RACHEL M. CHALMERS ◽  
GUY ROBINSON ◽  
EMILY HOTCHKISS ◽  
CLAIRE ALEXANDER ◽  
SOPHIE MAY ◽  
...  
Keyword(s):  
Cryptosporidium Parvum ◽  
Tandem Repeats ◽  
Repeat Unit ◽  
Unit Length ◽  
Variable Number ◽  
Multiple Locus ◽  
Chain Reactions ◽  
Outbreak Investigations ◽  
Laboratory Surveillance

SUMMARYCryptosporidium parvum is the major cause of livestock and zoonotically-acquired human cryptosporidiosis. The ability to track sources of contamination and routes of transmission by further differentiation of isolates would assist risk assessment and outbreak investigations. Multiple-locus variable-number of tandem-repeats (VNTR) analysis provides a means for rapid characterization by fragment sizing and estimation of copy numbers, but structured, harmonized development has been lacking for Cryptosporidium spp. To investigate potential for application in C. parvum surveillance and outbreak investigations, we studied nine commonly used VNTR loci (MSA, MSD, MSF, MM5, MM18, MM19, MS9-Mallon, GP60 and TP14) for chromosome distribution, repeat unit length and heterogeneity, and flanking region proximity and conservation. To investigate performance in vitro, we compared these loci in 14 C. parvum samples by capillary electrophoresis in three laboratories. We found that many loci did not contain simple repeat units but were more complex, hindering calculations of repeat unit copy number for standardized reporting nomenclature. However, sequenced reference DNA enabled reproducible fragment sizing and inter-laboratory allele assignation based on size normalized to that of the sequenced fragments by both single round and nested polymerase chain reactions. Additional Cryptosporidium loci need to be identified and validated for robust inter-laboratory surveillance and outbreak investigations.

Epigenetic differences between male and female bovine blastocysts produced in vitro

2008 ◽  
Vol 32 (2) ◽  
pp. 264-272 ◽  
Author(s):  
P. Bermejo-Álvarez ◽  
D. Rizos ◽  
D. Rath ◽  
P. Lonergan ◽  
A. Gutierrez-Adan
Keyword(s):  
Tandem Repeats ◽  
Dna Methyltransferases ◽  
Blastocyst Stage ◽  
Variable Number ◽  
Mtdna Copy Number ◽  
Male And Female ◽  
Epigenetic Events ◽  
Binding Factor ◽  
The Difference

Epigenetic differences between male and female bovine blastocysts provide a plausible link between physiological and gene transcription differences observed between male and female embryos. The aim of this study was to examine sex-related epigenetic differences in bovine blastocysts produced in vitro. Oocytes were matured in vitro and inseminated with frozen-thawed sex-sorted (X or Y) and unsorted (control) bull sperm. Zygotes were cultured to blastocyst stage and were analyzed for embryo sexing, mtDNA content, telomere lengths, methylation analysis, and quantification of mRNA transcripts of DNA methyltransferases (Dnmt1, Dnmt3a, Dnmt3b) HMT1 hnRNP methyltransferase-like 2 (Hmt1), and interleukin enhancer binding factor 3 (Ilf3). There was a difference ( P < 0.05) in the mean mtDNA copy number between male (410,000 ± 23,000) and female (360,000 ± 21,000) blastocysts. Telomere length was shorter in male blastocysts ( P < 0.01). The level of methylation in a sequence near a variable number of tandem repeats minisatellite region [variable number of tandem repeats (VNTR)] in males (39.8% ± 4.8) was higher than in females (23.7% ± 3.1) ( P < 0.05); however, no differences were found in other regions analyzed. Moreover, transcription differences between sexes were observed for Dnmt3a, Dnmt3b, Hmt1, and Ilf3. These results provide evidence of epigenetic differences between male and female bovine in vitro produced embryos and suggest that before initiation of gonadal differentiation, epigenetic events may modulate the difference between speed of development, metabolism, and transcription observed during preimplantation development between male and female embryos.

scholarly journals Intragenomic heterogeneity of intergenic ribosomal DNA spacers in Cucurbita moschata is determined by DNA minisatellites with variable potential to form non-canonical DNA conformations

DNA Research ◽  
2019 ◽  
Vol 26 (3) ◽  
pp. 273-286 ◽  
Author(s):  
Roman Matyášek ◽  
Alena Kuderová ◽  
Eva Kutílková ◽  
Marek Kučera ◽  
Aleš Kovařík
Keyword(s):  
Tandem Repeats ◽  
Intergenic Spacer ◽  
Structural Diversity ◽  
Structural Heterogeneity ◽  
Variable Number ◽  
Mutual Arrangement ◽  
Cucurbita Moschata ◽  
Sequence Motifs ◽  
Dna Conformations ◽  
High Selection

Abstract The intergenic spacer (IGS) of rDNA is frequently built of long blocks of tandem repeats. To estimate the intragenomic variability of such knotty regions, we employed PacBio sequencing of the Cucurbita moschata genome, in which thousands of rDNA copies are distributed across a number of loci. The rRNA coding regions are highly conserved, indicating intensive interlocus homogenization and/or high selection pressure. However, the IGS exhibits high intragenomic structural diversity. Two repeated blocks, R1 (300–1250 bp) and R2 (290–643 bp), account for most of the IGS variation. They exhibit minisatellite-like features built of multiple periodically spaced short GC-rich sequence motifs with the potential to adopt non-canonical DNA conformations, G-quadruplex-folded and left-handed Z-DNA. The mutual arrangement of these motifs can be used to classify IGS variants into five structural families. Subtle polymorphisms exist within each family due to a variable number of repeats, suggesting the coexistence of an enormous number of IGS variants. The substantial length and structural heterogeneity of IGS minisatellites suggests that the tempo of their divergence exceeds the tempo of the homogenization of rDNA arrays. As frequently occurring among plants, we hypothesize that their instability may influence transcription regulation and/or destabilize rDNA units, possibly spreading them across the genome.

scholarly journals Threonine-Rich Repeats Increase Fibronectin Binding in the Candida albicans Adhesin Als5p

2006 ◽  
Vol 5 (10) ◽  
pp. 1664-1673 ◽  
Author(s):  
Jason M. Rauceo ◽  
Richard De Armond ◽  
Henry Otoo ◽  
Peter C. Kahn ◽  
Stephen A. Klotz ◽  
...  
Keyword(s):  
Candida Albicans ◽  
Tandem Repeats ◽  
Surface Display ◽  
Cell Aggregation ◽  
Cellular Adhesion ◽  
Variable Number ◽  
Fungal Cell ◽  
T Domain ◽  
Α Helix

ABSTRACT Commensal and pathogenic states of Candida albicans depend on cell surface-expressed adhesins, including those of the Als family. Mature Als proteins consist of a 300-residue N-terminal region predicted to have an immunoglobulin (Ig)-like fold, a 104-residue conserved Thr-rich region (T), a central domain of a variable number of tandem repeats (TR) of a 36-residue Thr-rich sequence, and a heavily glycosylated C-terminal Ser/Thr-rich stalk region, also of variable length (N. K. Gaur and S. A. Klotz, Infect. Immun. 65: 5289-5294, 1997). Domain deletions in ALS5 were expressed in Saccharomyces cerevisiae to excrete soluble protein and for surface display. Far UV circular dichroism indicated that soluble Ig-T showed a single negative peak at 212 nm, consistent with previous data indicating that this region has high β-sheet content with very little α-helix. A truncation of Als5p with six tandem repeats (Ig-T-TR6) gave spectra with additional negative ellipticity at 200 nm and, at 227 to 240 nm, spectra characteristic of a structure with a similar fraction ofβ -sheet but with additional structural elements as well. Soluble Als5p Ig-T and Ig-T-TR6 fragments bound to fibronectin in vitro, but the inclusion of the TR region substantially increased affinity. Cellular adhesion assays with S. cerevisiae showed that the Ig-T domain mediated adherence to fibronectin and that TR repeats greatly increased cell-to-cell aggregation. Thus, the TR region of Als5p modulated the structure of the Ig-T region, augmented cell adhesion activity through increased binding to mammalian ligands, and simultaneously promoted fungal cell-cell interactions.

scholarly journals In vivo passage of Salmonella Typhimurium results in minor mutations in the bacterial genome and increases in vitro invasiveness

2019 ◽  
Vol 50 (1) ◽  
Author(s):  
Andrea R. McWhorter ◽  
Rick Tearle ◽  
Talia S. Moyle ◽  
Kapil K. Chousalkar
Keyword(s):  
Parent Strain ◽  
Tandem Repeats ◽  
Bacterial Genome ◽  
Variable Number ◽  
Single Infection ◽  
Typing Method ◽  
Mlva Type ◽  
Layer Hens

Abstract Eggs and raw or undercooked egg-containing food items are frequently identified as the bacterial source during epidemiolocal investigation of Salmonella outbreaks. Multi-locus variable number of tandem repeats analysis (MLVA) is a widely used Salmonella typing method enabling the study of diversity within populations of the same serotype. In vivo passage, however, has been linked with changes in MLVA type and more broadly the Salmonella genome. We sought to investigate whether in vivo passage through layer hens had an effect on MLVA type as well as the bacterial genome and whether any mutations affected bacterial virulence. Layer hens were infected with either Salmonella Typhimurium DT9 (03-24-11-11-523) as part of a single infection or were co-infected with an equal amount of Salmonella Mbandaka. Salmonella shedding in both single and co-infected birds was variable over the course of the 16-week experiment. Salmonella Typhimurium and Salmonella Mbandaka were identified in feces of co-infected birds. Salmonella colonies isolated from fecal samples were subtyped using MLVA. A single change in SSTR-6 was observed in Salmonella Typhimurium strains isolated from co-infected birds. Isolates of Salmonella Typhimurium of both the parent (03-24-11-11-523) and modified (03-24-12-11-523) MLVA type were sequenced and compared with the genome of the parent strain. Sequence analysis revealed that in vivo passaging resulted in minor mutation events. Passaged isolates exhibited significantly higher invasiveness in cultured human intestinal epithelial cells than the parent strain. The microevolution observed in this study suggests that changes in MLVA may arise more commonly and may have clinical significance.

Quantitative evaluation of post-bone marrow transplant engraftment status using fluorescent-labeled variable number of tandem repeats

2000 ◽  
Vol 05 (2) ◽  
pp. 129-138
Author(s):  
Robert A. Luhm ◽  
Daniel B. Bellissimo ◽  
Arejas J. Uzgiris ◽  
William R. Drobyski ◽  
Martin J. Hessner
Keyword(s):  
Bone Marrow ◽  
Bone Marrow Transplant ◽  
Quantitative Evaluation ◽  
Tandem Repeats ◽  
Marrow Transplant ◽  
Variable Number

Healing effects of curcumin nanoparticles in deep tissue injury mouse model.

2020 ◽  
Vol 18 ◽  
Author(s):  
Zirui Zhang ◽  
Shangcong Han ◽  
Panpan Liu ◽  
Xu Yang ◽  
Jing Han ◽  
...  
Keyword(s):  
Wound Healing ◽  
Mrna Expression ◽  
Tissue Injury ◽  
Inflammatory Cells ◽  
Pcr Analysis ◽  
Protective Effects ◽  
Deep Tissue ◽  
Deep Tissue Injury

Background: Chronic inflammation and lack of angiogenesis are the important pathological mechanisms in deep tissue injury (DTI). Curcumin is a well-known anti-inflammatory and antioxidant agent. However, curcumin is unstable under acidic and alkaline conditions, and can be rapidly metabolized and excreted in the bile, which shortens its bioactivity and efficacy. Objective: This study aimed to prepare curcumin-loaded poly (lactic-co-glycolic acid) nanoparticles (CPNPs) and to elucidate the protective effects and underlying mechanisms of wound healing in DTI models. Methods: CPNPs were evaluated for particle size, biocompatibility, in vitro drug release and their effect on in vivo wound healing. Results : The results of in vivo wound closure analysis revealed that CPNP treatments significantly improved wound contraction rates (p<0.01) at a faster rate than other three treatment groups. H&E staining revealed that CPNP treatments resulted in complete epithelialization and thick granulation tissue formation, whereas control groups resulted in a lack of compact epithelialization and persistence of inflammatory cells within the wound sites. Quantitative real-time PCR analysis showed that treatment with CPNPs suppressed IL-6 and TNF-α mRNA expression, and up-regulated TGF-β, VEGF-A and IL-10 mRNA expression. Western blot analysis showed up-regulated protein expression of TGF-β, VEGF-A and phosphorylatedSTAT3. Conclusion: Our results showed that CPNPs enhanced wound healing in DTI models, through modulation of the JAK2/STAT3 signalling pathway and subsequent upregulation of pro-healing factors.

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