Dimeric and trimeric fusion proteins generated with fimbrial adhesins of uropathogenic Escherichia coli

Urinary tract infections (UTIs) are associated with high rates of morbidity and mortality worldwide, and uropathogenic Escherichia coli (UPEC) is the main etiologic agent. Fimbriae assembled on the bacterial surface are essential for adhesion in the urinary tract epithelium. In this study, the FimH, CsgA, and PapG adhesins were fused to generate biomolecules as potential target vaccines against UTIs. The fusion protein design was generated using bioinformatics tools, and template fusion gene sequences were synthesized by GenScript in the following order fimH-csgA-papG-fimH-csgA (fcpfc) linked to the nucleotide sequence encoding the EAAAK5 peptide. Monomeric (fimH, csgA, and papG), dimeric (fimH-csgA), and trimeric (fimH-csgA-papG) genes were cloned into the pLATE31 expression vector and generated products of 1040, 539, 1139, 1442, and 2444 bp, respectively. Fusion protein expression in BL21 E. coli was induced with 1 mM IPTG, and His-tagged fusion proteins were purified under denaturing conditions and refolded by dialysis using C-buffer. Coomassie blue-stained SDS-PAGE gels and Western blot analysis revealed bands of 29.5, 11.9, 33.9, 44.9, and 82.1 kDa corresponding to FimH, CsgA, PapG, FC, and FCP proteins, respectively. Mass spectrometry analysis by MALDI-TOF/TOF revealed specific peptides that confirmed the fusion protein structures. Dynamic light scattering analysis revealed the polydispersed state of the fusion proteins. The FimH, CsgA, and PapG stimulated the release of 372 to 398 pg/mL IL-6; interestingly, the FC and FCP stimulated the release of 464.79 pg/mL (p ≤ 0.018) and 521.24 pg/mL (p ≤ 0.002) IL-6, respectively. In addition, the FC and FCP stimulated the release of 398.52 pg/mL (p ≤ 0.001) and 450.40 pg/mL (p ≤ 0.002) IL-8, respectively. High levels of IgA and IgG antibodies in human sera reacted against the fusion proteins, and under identical conditions, low levels of IgA and IgG antibodies were detected in human urine. Rabbit polyclonal antibodies generated against FimH, CsgA, PapG, FC, and FCP blocked the adhesion of the CFT073 E. coli strain to HTB5 bladder cells. In conclusion, the FC and FCP proteins were highly stable, demonstrated antigenic properties, and induced cytokine release (IL-6 and IL-8); furthermore, antibodies generated against these proteins showed protection against bacterial adhesion

Correction: Features of urinary Escherichia coli isolated from children with complicated and uncomplicated urinary tract infections in Mexico.

[This corrects the article DOI: 10.1371/journal.pone.0204934.]

Features of urinary Escherichia coli isolated from children with complicated and uncomplicated urinary tract infections in Mexico.

The Hospital Infantil de México Federico Gómez (HIMFG) is a tertiary care hospital in Mexico City where Escherichia coli is frequently isolated from the urine samples of pediatric patients with urinary tract infections. A collection of 178 urinary Escherichia coli (UEc) isolates associated with complicated and uncomplicated urinary tract infections were evaluated in this study. The patterns of resistance to 9 antibiotic classes showed that 60.7% of the UEc isolates had a highly multidrug-resistant (MDR) profile. Genetic diversity analyses of the UEc isolates showed a high variability and revealed 16 clusters associated with four phylogenetic groups, namely, groups A, B1, B2, and D. Phylogenetic group B2 was widely associated with the 16 clusters as well as with virulence and fitness genes. The virulence and fitness genes in the UEc isolates, which included fimbriae-, siderophore-, toxin-, and mobility-associated genes, were grouped as occurring at a low, variable, or high frequency. Interestingly, only the papF gene could be amplified from some UEc isolates, and the sequence analysis of the pap operon identified an insertion sequence (IS) element and gene loss. These data suggested pathoadaptability and the development of immune system evasion, which was confirmed by the loss of P fimbriae-associated agglutination in the UEc isolates. E. coli clone O25-ST131 had a prevalence of 20.2% among the UEc isolates; these isolates displayed both a highly MDR profile and the presence of the papGII, fimH, papGIII, iutD, sat, hlyA, and motA genes. In conclusion, the UEc isolates from complicated urinary tract infection (cUTI) were characterized as being MDR, highly genetically diverse, and associated with phylogenetic group B2 and many virulence and fitness genes. Additionally, gene loss and IS elements were identified in some UEc isolates identified as clone O25-ST131

[Antibiotic prescribing in respiratory tract infections and predictive factors for their use]

ResumenObjetivoEl estudio Happy Audit es un proyecto financiado por la Comunidad Europea, y tiene por objetivo mejorar la prescripción antibiótica en las infecciones del tracto respiratorio (ITR). Se realizó el presente estudio para conocer el tratamiento antibiótico administrado en las ITR en España y conocer qué criterios se asocian a su prescripción.DiseñoEstudio transversal realizado en enero y febrero de 2008.EmplazamientoCentros de atención primaria.ParticipantesMédicos de atención primaria de España que registraron durante 3 semanas todas las ITR mediante una plantilla.Mediciones principalesEdad y sexo, días con síntomas, signos presentes (fiebre, tos, otorrea purulenta, odinofagia, exudado amigdalar, adenopatías cervicales, disnea, aumento de esputo, esputo purulento), diagnóstico, tratamiento antibiótico y demanda de antibiótico. Se efectuó regresión logística y se consideró como variable dependiente la prescripción antibiótica.ResultadosDe los 332 médicos a los que se invitó a participar, 309 (93,1%) rellenaron los registros. Se registraron 16.751 ITR; las más frecuentes fueron el resfriado (39,7%), la faringitis (14,4%) y la bronquitis aguda (12,6%). Se prescribieron antibióticos en 4.675 ITR (27,9%), principalmente en neumonía (89,9%), amigdalitis (88,9%) y otitis media (87,3%). El criterio que más se asoció a prescripción antibiótica fue la presencia de exudado amigdalar (odds ratio [OR] de 32,1; intervalo de confianza [IC] del 95%: 24,5 a 42), seguido de otorrea (OR de 25,2; IC del 95%: 18,2 a 35) y esputo purulento (OR de 18,1; IC del 95%: 15,5 a 21,2); en cambio, fue protectora la tos (OR de 0,4; IC del 95%: 0,3 a 0,5).DiscusiónLa prescripción antibiótica en las ITR es muy elevada en España. Este estudio aporta información sobre factores predictores de tratamiento antibiótico fundamental para promover un uso más racional de antibióticos.AbstractObjectiveHappy Audit project is a European-funded survey aimed at reducing antibiotic prescribing for respiratory tract infections (RTI). The aim of this study is to investigate the antibiotic treatment administered for these RTIs in Spain and to find out which criteria are associated with its use.DesignCross-sectional study carried out in January and February 2008.SettingPrimary health care.ParticipantsGeneral practitioners registered all the RTI during a 3-week period using a template.Principal measurementsAge and gender, days with symptoms, signs presented (fever, cough, purulent ear discharge, sore throat, tonsillar exudate, swollen neck glands, dyspnoea, increase in sputum, purulent sputum), diagnosis, antibiotic therapy and demand of antibiotics. A logistic regression with the prescription of antibiotic as the dependent variable was performed.ResultsOut of the 332 physicians invited to participate, 309 filled in and returned the templates (93.1%), registering 16,751 RTIs, with the common cold (39.7%), pharyngitis (14.4%) and acute bronchitis (12.6%) being the most common. Antibiotic therapy was given to 4,675 RTIs (27.9%), mainly for pneumonia (89.9%), tonsillitis (88.9%), and otitis media (87.3%). The criterion most associated with antibiotic therapy was the presence of tonsillar exudate (OR: 32.1; 95CI%: 24.5–42), followed by ear discharge (25.2; 95%CI: 18.2–35) and purulence of sputum (18.1; 95%CI: 15.5–21.2); conversely, cough (OR: 0.4; 95%CI: 0.3–0.5) was considered as protective factor.DiscussionAntibiotic treatment for RTIs is very high in our country. This study provides information on the criteria that predict this antibiotic therapy and is important to take into account if a more rational use of antibiotics is required

COVID-19 in hospitalized HIV-positive and HIV-negative patients : A matched study

CatedresObjectives: We compared the characteristics and clinical outcomes of hospitalized individuals with COVID-19 with [people with HIV (PWH)] and without (non-PWH) HIV co-infection in Spain during the first wave of the pandemic. Methods: This was a retrospective matched cohort study. People with HIV were identified by reviewing clinical records and laboratory registries of 10 922 patients in active-follow-up within the Spanish HIV Research Network (CoRIS) up to 30 June 2020. Each hospitalized PWH was matched with five non-PWH of the same age and sex randomly selected from COVID-19@Spain, a multicentre cohort of 4035 patients hospitalized with confirmed COVID-19. The main outcome was all-cause in-hospital mortality. Results: Forty-five PWH with PCR-confirmed COVID-19 were identified in CoRIS, 21 of whom were hospitalized. A total of 105 age/sex-matched controls were selected from the COVID-19@Spain cohort. The median age in both groups was 53 (Q1-Q3, 46-56) years, and 90.5% were men. In PWH, 19.1% were injecting drug users, 95.2% were on antiretroviral therapy, 94.4% had HIV-RNA < 50 copies/mL, and the median (Q1-Q3) CD4 count was 595 (349-798) cells/μL. No statistically significant differences were found between PWH and non-PWH in number of comorbidities, presenting signs and symptoms, laboratory parameters, radiology findings and severity scores on admission. Corticosteroids were administered to 33.3% and 27.4% of PWH and non-PWH, respectively (P = 0.580). Deaths during admission were documented in two (9.5%) PWH and 12 (11.4%) non-PWH (P = 0.800). Conclusions: Our findings suggest that well-controlled HIV infection does not modify the clinical presentation or worsen clinical outcomes of COVID-19 hospitalization

How do women living with HIV experience menopause? Menopausal symptoms, anxiety and depression according to reproductive age in a multicenter cohort

CatedresBackground: To estimate the prevalence and severity of menopausal symptoms and anxiety/depression and to assess the differences according to menopausal status among women living with HIV aged 45-60 years from the cohort of Spanish HIV/AIDS Research Network (CoRIS). Methods: Women were interviewed by phone between September 2017 and December 2018 to determine whether they had experienced menopausal symptoms and anxiety/depression. The Menopause Rating Scale was used to evaluate the prevalence and severity of symptoms related to menopause in three subscales: somatic, psychologic and urogenital; and the 4-item Patient Health Questionnaire was used for anxiety/depression. Logistic regression models were used to estimate odds ratios (ORs) of association between menopausal status, and other potential risk factors, the presence and severity of somatic, psychological and urogenital symptoms and of anxiety/depression. Results: Of 251 women included, 137 (54.6%) were post-, 70 (27.9%) peri- and 44 (17.5%) pre-menopausal, respectively. Median age of onset menopause was 48 years (IQR 45-50). The proportions of pre-, peri- and post-menopausal women who had experienced any menopausal symptoms were 45.5%, 60.0% and 66.4%, respectively. Both peri- and post-menopause were associated with a higher likelihood of having somatic symptoms (aOR 3.01; 95% CI 1.38-6.55 and 2.63; 1.44-4.81, respectively), while post-menopause increased the likelihood of having psychological (2.16; 1.13-4.14) and urogenital symptoms (2.54; 1.42-4.85). By other hand, post-menopausal women had a statistically significant five-fold increase in the likelihood of presenting severe urogenital symptoms than pre-menopausal women (4.90; 1.74-13.84). No significant differences by menopausal status were found for anxiety/depression. Joint/muscle problems, exhaustion and sleeping disorders were the most commonly reported symptoms among all women. Differences in the prevalences of vaginal dryness (p = 0.002), joint/muscle complaints (p = 0.032), and sweating/flush (p = 0.032) were found among the three groups. Conclusions: Women living with HIV experienced a wide variety of menopausal symptoms, some of them initiated before women had any menstrual irregularity. We found a higher likelihood of somatic symptoms in peri- and post-menopausal women, while a higher likelihood of psychological and urogenital symptoms was found in post-menopausal women. Most somatic symptoms were of low or moderate severity, probably due to the good clinical and immunological situation of these women