Antibiotic resistance and virulence potentials of E. faecalis and E. faecium in hospital wastewater: a case study in Ardabil, Iran.

Hospital wastewater can contaminate the environment with antibiotic-resistant and virulent bacteria. We analyzed wastewater samples from four hospitals in Ardabil province, Iran for Enterococcus faecium and Enterococcus faecalis using culture and molecular methods. We also performed antimicrobial susceptibility testing and polymerase chain reaction testing for resistance and virulence genes. Out of 141 enterococci isolates, 68.8% were E. faecium and 23.4% were E. faecalis. Ciprofloxacin and rifampicin showed the highest level of resistance against E. faecalis and E. faecium isolates at 65%. High-level gentamicin resistance (HLGR), high-level streptomycin resistance (HLSR), ampicillin, and vancomycin resistance were observed in 25, 5, 10, and 5.15% of E. faecium, and 15, 6, 15, and 3.03% of E. faecalis isolates, respectively. The ant(6')-Ia and ant(3')-Ia genes that were responsible for streptomycin resistance were observed in HLSR isolates and aph(3')-IIIa and aac(6') Ie-aph(2″)-Ia genes accounting for gentamicin resistance were detected in HLGR isolates. vanA was the predominant gene detected in vancomycin-resistant isolates. The majority of isolates were positive for gelE, asa1, esp, cylA, and hyl virulence genes. We found that drug-resistant and virulent E. faecalis and E. faecium isolates were prevalent in hospital wastewater. Proper treatment strategies are required to prevent their dissemination into the environment.

Faecal carriage of high-level aminoglycoside-resistant and ampicillin-resistant Enterococcus species in healthy Iranian children.

OBJECTIVES: High-level aminoglycoside, ampicillin and vancomycin resistance and virulence genes among enterococcal isolates collected from healthy middle-school children in Ardabil, Iran, during 2016 were investigated. METHODS: Totally, 305 faecal specimens were collected. Isolates underwent antimicrobial susceptibility testing, virulence gene detection and molecular typing. RESULTS: Totally, 409 enterococcal isolates were collected, comprising Enterococcus faecium (235; 57.5%), Enterococcus faecalis (56; 13.7%) and other Enterococcus spp. (118; 28.9%). Overall, 71 (17.4%), 11 (2.7%) and 10 (2.4%) isolates were identified as high-level streptomycin-resistant (HLSR), high-level gentamicin-resistant (HLGR) and ampicillin-resistant (AR), respectively. Among HLSR isolates, 40 (56.3%), 5 (7.0%) and 26 (36.6%) were E. faecium, E. faecalis and other Enterococcus spp., respectively. Among HLGR isolates 4 (36.4%) and 7 (63.6%) and among AR isolates 7 (70.0%) and 3 (30.0%) were E. faecium and other Enterococcus spp., respectively. Accordingly, 21.6%, 3.6% and 3.3% of subjects were colonised with HLSR, HLGR and AR Enterococcus spp. Carriage of HLGR, HLSR and AR isolates was associated with prior antibiotic consumption (P≤0.05). Additionally, male sex and antacid consumption were associated with AR enterococcal carriage. Moreover, 69 (97.2%), 10 (90.9%) and 9 (90.0%) of HLSR, HLGR and AR isolates were multidrug-resistant, respectively. No vancomycin-resistant enterococci were detected. ERIC-PCR revealed high genetic diversity among isolates. gelE and asa1 were major virulence genes both in E. faecalis and E. faecium. Presence of gelE was associated with HLSR and HLGR phenotypes (P≤0.05). CONCLUSION: Community intestinal carriage of HLSR enterococci was high; however, carriage of HLGR and AR enterococci was low.

Use of a bacterially expressed human factor IX light chain to develop polyclonal antibody anti-hFIX.

Hemophilia B is an X-linked recessive bleeding disorder caused by deficiency or malfunctioning of human coagulation factor IX (hFIX). Hemophilia B patients are treated at present by infusion of plasma derived hFIX which is not always efficient, because development of anti-hFIX antibodies (alloantibodies) in some cases inhibits the activity of the infused hFIX. The hFIX alloantibodies are directed against gamma-carboxyglutamic acid residues (Gla-domain) or protease domain in hFIX light chain. An epitope-containing fragment of hFIX light-chain was expressed in a T7-based Escherichia coli expression system and after purification, it was used for the immunization of rabbit to develop specific antibodies anti-hFIX. The plasma, derived from the immunized rabbit, was shown to be able to detect the normal hFIX, which indicates for the presence of a specific anti-hFIX antibody and supporting that a bacterially expressed hFIX subfragment might be able to neutralize the alloantibodies. Considering the importance of hFIX and its related investigations, both the produced hFIX antigen and its corresponding antibody will play important roles for experiments dealing with the production of hFIX and studies involved in the neutralization of the hFIX inhibitors in hFIX-related disorders and other clinical applications.