Molecular characteristics of extended-spectrum β-lactamase-producing Escherichia coli from Rio de Janeiro,Brazil

Twenty-five extended-spectrum β-lactamase (ESBL)-producing Escherichia coli clinical isolates from Rio de Janeiro, Brazil were characterized by isoelectric focusing, PCR and sequencing of bla^ESBL genes, plasmid-mediated quinolone resistance determinants, phylogenetic groups, replicon typing, pulsed-field electrophoresis, and multilocus sequencing typing. Twenty-three (92%) ESBL-producing E. coli isolates were positive for bla^CTX-M genes, aac(6′)-Ib-cr, and qnrB. Genetic relatedness of ESBL producers clustered seven (28%) CTX-M-15-producing isolates as sequence type (ST)410, clonal complex (CC) 23, and two (8%) as clone O25-ST131. Our results illustrate the predominance of phylogroup A (52%), ST410 (CC 23) and CTX-M-15 among ESBL-producing E. coli isolates from hospitals in Rio de Janeiro.     

Virulence markers of enteroaggregative Escherichia coli isolated from children and adults with diarrhea in Brasília,Brazil

Escherichia coli strains isolated from sporadic cases of acute diarrhea in children and adults and from children without diarrhea were investigated for the presence of the pAA plasmid. Strains harboring the pAA plasmid were isolated at similar frequencies from children with (19.6%) and without (10.8%) diarrhea and from adults with diarrhea (11.8%). The genotypic and phenotypic virulence markers of these strains were further analyzed. Most of the strains were positive for EAST1 (73%), and this toxin was detected significantly more frequently in strains from children with diarrhea than in strains from adults with diarrhea (P < 0.05). Likewise, pic sequences were detected significantly more frequently in strains from children with diarrhea than in strains from adults with diarrhea (P < 0.005) and controls (P < 0.025). Furthermore, the association of pAA positivity (pAA(+)) and pic positivity (pic(+)) was more frequently found for strains from children with diarrhea than for strains from controls, indicating that pAA(+) pic(+) strains may represent a subset of pAA(+) strains associated with disease in children. Most of the strains (82.5%) adhered to cells presenting the typical aggregative pattern. The frequency of occurrence of enteropathogenic E. coli (EPEC) serogroups in the strains from children with diarrhea was very high (56%), while none of the strains from adults with diarrhea belonged to EPEC serogroups. Extraintestinal virulence markers were very commonly found in strains from adults with diarrhea. The frequencies of occurrence of the adhesins AFA and SFA were significantly higher in strains from adults with diarrhea than in strains from children with diarrhea. More than one extraintestinal virulence marker was found in 58% of the strains from adults with diarrhea but in only 7.7% of the strains from children with diarrhea. Our results show that pAA(+) strains isolated from children and adults with diarrhea present very different profiles when enteroaggregative E. coli virulence markers, serotypes, and extraintestinal virulence markers are considered.     

E. coli: resistance to quinolones and beta-lactams of clinical strains isolated in the Franche-Comté region of France

AIM OF THE STUDY: Numerous European studies have reported an increase of resistance to quinolones among E. coli. We conducted a regional study to update our knowledge on this evolution. MATERIALS AND METHODS: We evaluated the resistance phenotype and genotype of 115 clinical strains of E. coli. We collected data on individual treatment with fluoroquinolones, and the evolution of the use of these antimicrobial agents. RESULTS: Resistance to nalidixic acid and ciprofloxacin was 13.0 and 6.9, respectively. The frequency of resistance increased from 1999 to 2001, from 7.5% to 13.0% for nalidixic acid and from 5.4% to 6.9% for fluoroquinolones. Resistance to quinolones was significantly associated to beta-lactams resistance and was slightly higher for nosocomial isolates compared to community-acquired isolates. Previous treatment with fluoroquinolones was the major risk factor associated to E. coli resistance. From 1997 to 2001, fluoroquinolones use has increased in our hospital and particularly in the community. Analysis of molecular epidemiology shows a large clonal diversity among E. coli isolates. CONCLUSION: This study confirms the evolution through resistance to quinolones of E. coli isolates. This observation is not due to dissemination of resistant clonal strains and the selective pressure exerted by fluoroquinolones influences this evolution. Therapeutic alternatives, surveillance, and restriction of fluoroquinolones use are needed to control this spread of resistance.     

Identification of a high-risk heterosexual population for HIV prevention trials in Rio de Janeiro, Brazil. Projeto Praço Onze Study Group

The study of interventions to prevent HIV transmission requires access to populations with a high rate of HIV transmission. We estimated HIV incidence among heterosexual males and females who were seen at an HIV testing site in Rio de Janeiro, Brazil. Stored sera from individuals who visited the site between March and December 1998 were analyzed using the sensitive/less sensitive (S/LS) assay and a chart abstraction was performed. During the study period, 6353 serum samples were tested. Of those tested, 1203 were found to be HIV-seropositive or indeterminate, of which 1050 (87%) remained available for further testing. In addition, 84 serum samples, representing 63 adults, were found to produce results suggesting early HIV infection. Of these, 14 were heterosexual and female (median age, 38 years), and 19 were heterosexual and male (median age, 25 years). The estimated HIV seroincidence was 1.9 (95% confidence limits (CL), 0.9%-3.9%) and 2.8 (95% CL, 1.4%-5.3%) per 100 person-years among heterosexual women and men, respectively. A survey on willingness to participate in future placebo-controlled HIV vaccine trials in this population indicated that 54.5% and 53.9% of heterosexual women and men, respectively, indicated that they would definitely be willing to participate. We have identified a heterosexual population in Rio de Janeiro with a high rate of HIV transmission willing to participate in placebo-controlled vaccine trials. This study demonstrates the usefulness of the newly described S/LS assay, which allows one to estimate HIV incidence from single serum specimens.     

The characteristics of metallo-β-lactamase-producing gram-negative bacilli isolated from sputum and urine: a single center experience in Korea

Metallo-β-lactamase (MBL) production usually results in high-level resistance to most β-lactams, and a rapid spread of MBL producing major gram-negative pathogens is a matter of particular concern worldwide. However, clinical data are scarce and most studies compared MBL producer (MP) with MBL non-producer (MNP) strains which included carbapenem susceptible isolates. Therefore, we collected clinical data of patients in whom imipenem-nonsusceptible Pseudomonas aeruginosa (PA) and Acinetobacter baumannii (AB) were isolated from sputum or urine, and investigated MBL production and the risk factors related with MBL acquisition. The antimicrobial susceptibility patterns were also compared between MPs and imipenem-nonsusceptible MNPs (INMNP). Among the 176 imipenem-nonsusceptible isolates, 12 MPs (6.8%) were identified. There was no identifiable risk factor that contributed to the acquisition of MPs when compared to INMNPs, and case-fatalities were not different between the two groups. The percentage of susceptible isolates was higher among MPs for piperacilin/tazobactam and fluoroquinolones while that of ceftazidime was higher in INMNPs (p < 0.05). As regards to aztreonam, which has been known to be a uniquely stable β-lactam against MBLs, susceptibility was preserved in only two isolates (16.7%) among MPs, and was not higher than that of INMNPs (23.2%). In conclusion, the contribution of MBLs to imipenem non-susceptibility in PA/ABs isolated from sputum and urine was relatively limited, and there was no significant risk factor associated with acquisition of MPs compared with INMNPs. However, limited susceptibility to aztreonam implies that MPs may hold additional resistance mechanisms, such as extended spectrum β-lactamases, AmpC β-lactamases, or other non-enzymatic mechanisms.     

The expression and preliminary evaluation of HPV6bL2△N360E7E6 fusion protein in E.coli for genital warts

Objective To express HPV6bL2△N360E7E6 fusion protein in E.coli and preliminarily evaluate its immune effect.Methods Three HPV6b gene fragments,which were L2(1-360 bp),E7 and E6,were fused by overlapping PCR,then were inserted into a prokaryotic expression vector and expressed in E.coli.C57BL/6 mice were immunized with purified fusion protein plus Al(OH)3 and/or CpG adjuvants through intramuscular route,the cellular and humoral immune responses were detected by IFN-γ ELISPOT and ELISA respectively.Results Protein plus CpG adjuvant could induce the strongest cellular immune response to E7 and E6,high antibody titer against L2 could be detected in all immunized groups but there were no significant difference among these groups.Conclutions HPV6bL2△N360E7E6 gene was successfully cloned into pQE30 vector and expressed in E.coli,the fusion protein was also purified and proved that could induce strong cellular and humoral immune responses with appropriate adjuvant in C57 BL/ 6 mice and could be used for future research.     

The expression and preliminary evaluation of HPV6bL2△N360E7E6 fusion protein in E.coli for genital warts

Objective To express HPV6bL2△N360E7E6 fusion protein in E.coli and preliminarily evaluate its immune effect.Methods Three HPV6b gene fragments,which were L2(1-360 bp),E7 and E6,were fused by overlapping PCR,then were inserted into a prokaryotic expression vector and expressed in E.coli.C57BL/6 mice were immunized with purified fusion protein plus Al(OH)3 and/or CpG adjuvants through intramuscular route,the cellular and humoral immune responses were detected by IFN-γ ELISPOT and ELISA respectively.Results Protein plus CpG adjuvant could induce the strongest cellular immune response to E7 and E6,high antibody titer against L2 could be detected in all immunized groups but there were no significant difference among these groups.Conclutions HPV6bL2△N360E7E6 gene was successfully cloned into pQE30 vector and expressed in E.coli,the fusion protein was also purified and proved that could induce strong cellular and humoral immune responses with appropriate adjuvant in C57 BL/ 6 mice and could be used for future research.     

The genetic and antigenic diversity of avian influenza viruses isolated from domestic ducks, muscovy ducks, and chickens in northern and southern Vietnam, 2010–2012

To estimate the prevalence of avian influenza virus infection in Vietnam, surveillance was conducted in domestic and wild birds from households, live-bird markets, slaughtering sites, and bird sanctuaries in Vietnam between October 2010 and October 2012. Of the 4,550 samples collected, 226 influenza A virus isolates were obtained from domestic ducks, muscovy ducks, and chickens. Of these, 25 and 22 H5N1 highly pathogenic avian influenza viruses (HPAIVs) were isolated from apparently healthy domestic ducks in live-bird markets and slaughtering sites in northern and southern Vietnam, respectively. The HA genes of H5 viruses isolated from birds in northern Vietnam phylogenetically belonged to the genetic clade 2.3.2.1 and those in southern Vietnam belonged to the genetic clade 1.1. In addition, 39 H3, 12 H4, 1 H5, 93 H6, 2 H7, 18 H9, 3 H10, and 11 H11 viruses were isolated. Phylogenetic and antigenic analyses of the H6 and H9 viruses revealed that they were closely related to the isolates obtained from domestic poultry in China. Phylogenetic analyses of internal gene segments of these isolates revealed that these viruses were circulating in both domestic and wild birds in Asia and reassortment events had occurred frequently. Therefore, it will be important to continue the surveillance and strict controls over the movement and trade of poultry and poultry products in order to eradicate H5N1 HPAIV from Asia.     

Influence of temperature,salinity and E. coli tissue content on immune gene expression in mussel: Results from a 2005–2008 survey

Several bivalves, including mussels, suffered from mortalities particularly in summer. To look for the possible effect of environmental parameters on immune capacities, Mytilus galloprovincialis were collected monthly from August 2005 to July 2008 from the Palavas Laguna, French Mediterranean coast. Q-PCR was used to quantify the expression of three antimicrobial peptide genes (defensin, mytilin B and myticin B), in addition to lysozyme and HSP70. House keeping gene was 28S rRNA. Defensin, myticin B and lysozyme appeared more expressed in spring–summer than in winter. In contrast, HSP70 expression was higher in winter. Statistical studies using principal component analysis (PCA) and multiple regression models revealed positive influence of temperature on 28S rRNA, defensin, myticin B and lysozyme expressions, but not on mytilin B and HSP70. The positive influence was significant for defensin and lysozyme expression, but relationships cannot be quantified. Similarly, salinity appeared to influence defensin expression, but this relationship cannot be quantified neither. E. coli tissue content appeared without influence. Consequently, there was no clear relationship between environmental parameters and immune-related gene expressions, demonstrating anti-infectious capabilities cannot be evaluated using only the expression of such genes as markers.     

A new species of Eimeria Schneider, 1875 from the Serra dos Órgãos National Park,Rio de Janeiro,Brazil, with notes on its endogenous development in the montane grass mouse,Akodon montensis Thomas, 1913 (Rodentia: Sigmodontinae)

Parasitology Research - A total of 53 specimens of the montane grass mouse, Akodon montensis Thomas, 1913 were collected in the Serra dos Órgãos National Park (SONP) in November 2014 and...