Detection and characterization of hybrid uropathogenic Escherichia coli strains among E. coli isolates causing community-acquired urinary tract infection.

Introduction. The main aetiological agent of urinary tract infection (UTI) is Escherichia coli, categorized as uropathogenic E. coli (UPEC). The genome of UPEC shows a high degree of plasticity, which leads to the emergence of 'intermediary strains' with different traits from the parental pathotypes.Gap Statement/Aim. We aimed to assess the frequency and types of the hybrid UPEC among isolates causing UTI and characterize virulence properties of these hybrid isolates molecularly and phenotypically.Methodology. After detection of intestinal pathogenic E. coli (IPEC) virulence markers among 200 UPEC isolates, they were assessed for the presence of 40 virulence genes (VGs) of extraintestinal, uropathogenic and diarrhoeagenic E. coli, phylogenetic group typing, phenotypic traits including biofilm formation, adherence and invasion to HeLa cells, haemolysis activity and antimicrobial resistance.Results. The analysis showed 21 (10.5 %) UPEC isolates carried enteroaggregative E. coli (EAEC) and enteropathogenic E. coli (EPEC) virulence markers. Twenty isolates carried the aggR (EAEC) and one the eae and escV genes (EPEC), which were classified as hybrid strains. The most commonly identified genes were fimH (71.5 %), fyuA (66.7 %), iutA (62 %), chuA (57.1) and traT (47.6 %). Biofilm production, adhesion and invasion were found among 17 (81), 18 (85.7) and 11 (52.4 %) hybrids, respectively. Investigation of the genetic characteristics, phylogenetic group and virulence profile of the detected hybrids revealed that they have genetic diversity and do not belong to a particular clonal lineage.Conclusion. The present study reveals that some UPEC may carry virulence markers of IPEC pathotypes. EAEC and EPEC seem to have a greater tendency to form hybrids and cause UTI. Further studies are needed to elucidate what factors contributed to survival in the urinary tract system and facilitate infection and whether these combinations lead to an increase in pathogenicity or not.

Expression, purification, and study on the efficiency of a new potent recombinant scFv antibody against the SARS-CoV-2 spike RBD in E. coli BL21.

Many efforts have been made around the world to combat SARS-CoV-2. Among these are recombinant antibodies considered to be suitable as an alternative for some diagnostics/therapeutics. Based on their importance, this study aimed to investigate the expression, purification, and efficiency of a new potent recombinant scFv in the E. coli BL21 (DE3) system. The expression studies were performed after confirming the scFv cloning into the pET28a vector using specific PCRs. After comprehensive expression studies, a suitable strategy was adopted to extract and purify periplasmic proteins using Ni2+-NTA resin. Besides the purified scFv, the crude bacterial lysate was also used to develop a sandwich ELISA (S-ELISA) for the detection of SARS-CoV-2. The use of PCR, E. coli expression system, western blotting (WB), and S-ELISA confirmed the functionality of this potent scFv. Moreover, the crude bacterial lysate also showed good potential for detecting SARS-CoV-2. This could be decreasing the costs and ease its utilization for large-scale applications. The production of high-quality recombinant proteins is essential for humankind. Moreover, with attention to the more aggressive nature of SARS-CoV-2 than other coronaviruses, the development of an effective detection method is urgent. Based on our knowledge, this study is one of the limited investigations in two fields: (1) The production of anti-SARS-CoV-2 scFv using E. coli [as a cheap heterologous host] in relatively high amounts and with good stability, and (2) Designing a sensitive S-ELISA for its detection. It may also be utilized as potent therapeutics after further investigations.

Delivery of antisense peptide nucleic acid by gold nanoparticles for the inhibition of virus replication.

Aim: We aim to use peptide nucleic acid (PNA) for antisense therapy against bovine viral diarrhea virus (BVDV), a surrogate model of human hepatitis C virus, and introduce an optimal approach for delivering PNA into the cell. Materials & methods: PNA was designed for hybridization to the 5'-untranslated region of BVDV RNA in order to form a heteroduplex structure and inhibit the translation and replication of virus. Gold nanoparticles (AuNPs) were used as a delivery system for PNA. Results: The cellular uptake of PNA-AuNPs and inhibition of BVDV infection in the middle stage of viral replication were found. Conclusion: Further research is warranted to develop AuNPs as a potential vehicle for delivering PNA in order to remove viruses from the infected cells.

In vitro anti-foot-and-mouth disease virus activity of magnesium oxide nanoparticles.

Foot-and-mouth disease (FMD) is an extremely contagious viral disease of cloven-hoofed animals that can lead to huge economic losses in the livestock production. No antiviral therapies are available for treating FMD virus (FMDV) infections in animals. The antiviral effects of magnesium oxide nanoparticles (MgO NPs) on the FMDV were investigated in cell culture. The viability of the cells after MgO NP treatment was determined using the MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] assay. The direct effects of MgO NPs on the FMDV in extracellular (virucidal assay) and also different stages of virus replication (antiviral assay) were evaluated by plaque reduction assay. The results showed that MgO NPs were safe at concentrations up to 250 µg/ml in the Razi Bovine kidney cell line. The treatments with NPs indicated that the MgO NPs exerted in vitro virucidal and antiviral activities. Plaque reduction assay revealed that MgO NPs can inhibit FMDV by more than 90% at the early stages of infection such as attachment and penetration but not after penetration. The results of this study suggested that NPs might be applied locally as an antiviral agent in early stages of infection in susceptible animals.

Biodesulfurization of dibenzothiophene by Gordonia sp. AHV-01 and optimization by using of response surface design procedure.

A novel desulfurizing bacterium has been isolated from oil-contaminated soils in Khuzestan. The ability for dibenzothiophene desulfurization and its biochemical pathway were investigated. The bacterium was identified as Gordonia sp. AHV-01 (Genbank Accession No HQ607780) by 16S rRNA gene sequencing. HPLC results and Gibb's assay were shown that dibenzothiophene desulfurized via 4S-pathway Maximum growth (0.426 g dry cells/L) and produced 2-hydroxybiphenyl (63.1 microM) were observed at 120 h of cultivation. By using of response surface design procedure the optimization of pH, temperature and rotary shaker round on the desulfurization reaction of isolate AHV-01 were performed. The optimum conditions were determined at pH of 7.0, temperature of 30 degrees C and rotary shaker round of 180 rpm. At these conditions, the dibenzothiophene desulfurization activity was increased and maximum 2-hydroxybiphenyl production was detected 70.29 microM at 96 h. According to these results, Isolate AHV-01 was capable to desulfurize dibenzothiophene via 4S-pathway and likely it can be useful to reduce organic sulfur contents of crude oil.

C-S targeted biodegradation of dibenzothiophene by Stenotrophomonas sp. NISOC-04.

Crude oil-contaminated soil samples were gathered across Khuzestan oilfields (National Iranian South Oil Company, NISOC) consequently experienced a screening procedure for isolating C-S targeted dibenzothiophene-biodegrading microorganisms with previously optimized techniques. Among the isolates, a bacterial strain was selected due to its capability of biodegrading dibenzothiophene in a C-S targeted manner in aqueous phases and medium mostly consisting of separately biphasic water-gasoline. The 16S rDNA of the isolate was amplified using eubacterial-specific primers and then sequenced. Based on sequence data analysis, the microorganism, designated NISOC-04, clustered most closely with the members of the genus Stenotrophomonas. Gas chromatography indicated that Stenotrophomonas sp. NISOC-04 utilizes 82% of starting 0.8 mM dibenzothiophene within a 48-h-long exponential growth phase. Growth curve analysis revealed the inability of Stenotrophomonas sp. NISOC-04 to utilize dibenzothiophene (DBT) as the exclusive carbon or carbon/sulfur source. Gibbs' assay showed no 2-hydroxy biphenyl accumulation, but HPLC confirmed the presence of 2-hydroxy biphenyl as the final product of DBT desulfurization. Under sulfur starvation, Stenotrophomonas sp. NISOC-04 produced a huge biomass with untraceable sulfur and utilized atmospheric insignificant sulfur levels.

Genotyping of the prevalent Chlamydia trachomatis strains involved in cervical infections in women in Ahvaz, Iran.

To determine the prevalence of cervical Chlamydia trachomatis genotypes in Iran for the first time and their association with three clinical symptoms/signs, i.e. abnormal vaginal discharge, lower abdominal pain (LAP) and swab-induced bleeding, and patient age, 620 cervical specimens were obtained from women with symptomatic genital infection referred to gynaecological clinics and 108 C. trachomatis-positive specimens were genotyped by direct omp1 gene PCR-RFLP analysis. Eight genotypes were identified. The most prevalent genotype was E (31.5 %), followed by F (23.1 %), D/Da (13 %), K (9.2 %), I (8.3 %), G (7.5 %), H (5.5 %) and J (1.9 %). For analysing the association of C. trachomatis genotypes with symptoms/signs and age, P-values were separately evaluated for genogroups and genotypes. The analysis of genogroups showed that women infected with genogroup F/G manifested the symptom of LAP significantly more often than those infected with the other genogroups (P=0.02), while the analysis of genotypes revealed that women infected with genotype F reported LAP slightly more often than women infected with the other genotypes (P=0.08). No significant correlation between genogroups and age was found; however, genotype E was somewhat less prevalent among women aged 25-34 years than among other age groups (P=0.08).

Utilization of dibenzothiophene as sulfur source by Microbacterium sp. NISOC-06.

Oil-polluted soils were sampled from National Iranian South Oil Company (NISOC) for isolation and screening of C-S and not C-C targeted Dibenzothiophene (DBT) degrading microorganisms. Microbacterium sp. NISOC-06, a C-S targeted DBT degrading bacterium, was selected and its desulfurization ability was studied in aqueous phase and water-gasoline biphasic systems. The 16srRNA gene was amplified using universal eubacteria-specific primers, PCR product was sequenced and the sequence of nearly 1,500 bp 16srDNA was studied. Based on Gas Chromatography results Microbacterium sp. NISOC-06 utilized 94.8% of 1 mM DBT during the 2 weeks of incubation. UV Spectrophotometry and biomass production measurements showed that the Microbacterium sp. NISOC-06 was not able to utilize DBT as a carbon source. There was no accumulation of phenolic compounds as Gibb's assay showed. Biomass production in a biphasic system for which DBT-enriched gasoline was used as the sulfur source indicated the capability of Microbacterium sp. NISOC-06 to desulfurize gasoline.