Nucleotide substitutions in the mexR, nalC and nalD regulator genes of the MexAB-OprM efflux pump are maintained in Pseudomonas aeruginosa genetic lineages.

Pseudomonas aeruginosa has different resistant mechanisms including the constitutive MexAB-OprM efflux pump. Single nucleotide polymorphisms (SNPs) in the mexR, nalC, and nalD repressors of this efflux pump can contribute to antimicrobial resistance; however, it is unknown whether these changes are mainly related to genetic lineages or environmental pressure. This study identifies SNPs in the mexR, nalC, and nalD genes in clinical and environmental isolates of P. aeruginosa (including high-risk clones). Ninety-one P. aeruginosa strains were classified according to their resistance to antibiotics, typified by multilocus sequencing, and mexR, nalC, and nalD genes sequenced for SNPs identification. The mexAB-oprM transcript expression was determined. The 96.7% of the strains were classified as multidrug resistant. Eight strains produced serine carbapenemases, and 11 strains metallo-ß-lactamases. Twenty-three new STs and high-risk clones ST111 and ST233 were identified. SNPs in the mexR, nalC, and nalD genes revealed 27 different haplotypes (patterns). Sixty-two mutational changes were identified, 13 non-synonymous. Haplotype 1 was the most frequent (n = 40), and mainly identified in strains ST1725 (33/40), with 57.5% pan drug resistant strains, 36.5% extensive drug resistant and two strains exhibiting serin-carbapenemases. Haplotype 12 (n = 9) was identified in ST233 and phylogenetically related STs, with 100% of the strains exhibiting XDR and 90% producing metallo-ß-lactamases. Haplotype 5 was highly associated with XDR and related to dead when compared to ST1725 and ST233 (RRR 23.34; p = 0.009 and RRR 32.01; p = 0.025). A significant relationship between the mexR-nalC-nalD haplotypes and phylogenetically related STs was observed, suggesting mutational changes in these repressors are highly maintained within genetic lineages. In addition, phylogenetically related STs showed similar resistant profiles; however, the resistance was (likely or partly) attributed to the MexAB-OprM efflux pump in 56% of the strains (only 45.05% showed mexA overtranscription), in the remaining strains the resistance could be attributed to carbapenemases or mechanisms including other pumps, since same SNPs in the repressor genes gave rise to different resistance profiles.

Inference from the analysis of genetic structure of Helicobacter pylori strains isolates from two paediatric patients with recurrent infection.

BACKGROUND: Helicobacter pylori recurrence after successful eradication is an important problem. Children are particularly vulnerable to reinfection, by intrafamilial transmission which facilitates the acquisition or recombination of new genetic information by this bacterium. We investigated the evolutionary dynamics of 80 H. pylori strains isolated from two paediatric patients with recurrent infection (recrudescence and reinfection). RESULTS: We characterized the virulence genes vacA (s1, m1, s2, and m2), cagA, cagE, and babA2 and performed multilocus sequence typing (MLST) on 7 housekeeping genes (atpA, efp, ureI, ppa, mutY, trpC, and yphC) to infer the evolutionary dynamics of the H. pylori strains through phylogenetic and genealogic inference analyses, genetic diversity analysis and the exploration of recombination events during recurrent infections. The virulence genotype vacAs1m1/cagA+/cagE+/babA2 was present at a high frequency, as were the EPIYA motifs EPIYA-A, -B and -C. Furthermore, the housekeeping genes of the H. pylori strains exhibited high genetic variation, comprising 26 new alleles and 17 new Sequence Type (ST). In addition, the hpEurope (76.5%) and hspWAfrica (23.5%) populations predominated among the paediatric strains. All strains, regardless of their ancestral affiliation, harboured western EPIYA motifs. CONCLUSIONS: This study provides evidence of the evolutionary dynamics of the H. pylori strains in two paediatric patients during recrudescence and reinfection events. In particular, our study shows that the strains changed during these events, as evidenced by the presence of different STs that emerged before and after treatment; these changes may be due to the accumulation of mutations and recombination events during the diversification process and recolonization of the patients by different genotypes.

Pediatric inguinal hernia repair with a single-incision approach using an Endo Close™ suturing device.

BACKGROUND: Diverse techniques have been described for pediatric inguinal hernia repair, based on extraperitoneal [1-4] and intraperitoneal [5-8] methodologies. In this video, we describe a novel technique to repair pediatric inguinal hernia using an Endo Close™ suturing device by percutaneous puncture with a single incision. METHODS: With a transumbilical approach, a 5-mm trocar is inserted for a 30° laparoscope. A 3-mm incision is made, and the Endo Close™ suturing device (Covidien, Minneapolis, MN, USA), with a 2-0 polypropylene suture retained by the stylet, is inserted perpendicularly to the skin. An extraperitoneal dissection is made on a side the inguinal ring and the needle of the device penetrates the peritoneum through the inferior border. Then, the stylet mechanism is pushed to free the lasso inside the cavity. At the same incision site, the needle of the Endo Close™ is inserted again, but an extraperitoneal dissection is made on the other side of the ring, ensuring that the needle penetrates at the same exit orifice. Now, the suture lasso is recovered and retracted to close the ring. Finally, the suture is extracted and knots are tied extracorporeally at the level of the skin. RESULTS: A total of 34 patients (20 females and 14 males) underwent surgery with this procedure. Operative time for unilateral repair was 10-15 and 25-30 min for the bilateral repair (29 unilateral/5 bilateral). The patients experienced minimal postoperative pain. The follow-up period was 12 months with no complications, no recurrence and without cases of postoperative hydrocele. There were no injuries to the structures as vessels or vas deferens, and the esthetic outcome was excellent. CONCLUSIONS: The technique presents a simple, safe and reliable method to repair inguinal hernias in children. The long-term results of this novel technique will be evaluated in future studies.