Carvalho, Ana P.

Costa, Eduardo

Durão, Joana

Faustino, Margarida

Pereira, Ana Margarida

Pereira, Carla F.

Pereira, Joana Odila

Pintado, Manuela E.

Silva, Sara

English

Repositório Institucional da Universidade Católica Portuguesa

Effect of mannanoligosaccharides extracts in uropathogenic Escherichia coli adhesion in human bladder cellsMargarida Faustino a, Sara Silva a, Eduardo Costa a, Joana Odila Pereira a,b, Ana Margarida Pereira  a,b, Carla F. Pereira a, Joana Durão a,b , Manuela E. Pintado a,  Ana P. Carvalho aa Universidade Católica Portuguesa, CBQF - Centro de Biotecnologia e Química Fina – Laboratório Associado, Escola Superior de Biotecnologia, Rua Diogo Botelho 1327, 4169-005 Porto, Portugalb Amyris Bio Products Portugal Unipessoal Lda, PortugalIntroduction / Objectives MethodsConclusionsAcknowledgementsIn prophylaxys (Figure 3), 1 h of pretreatment appeared to be better than longer pre-exposure times, a likely consequence of extracts’ metabolization by either the cells or bacteria.The treatment did not result in any significant reduction of the pre-established infection. MOS extracts performed better in the competition one, as 2.5 mg/mL total mannoseallowed for a significant inhibition of UPEC in both extracts (MOS 1 (89.61± 1.37 %) and MOS 2 (79.78± 1.12 %). Regardless, the extracts demonstrated an interesting potential tobe used as prophylactic health supplements in the future.This work was financially supported by Amyris Bio Products Portugal Unipessoal Lda and Escola Superior de Biotecnologia – Universidade Católica Portuguesathrough Alchemy project- Capturing high value from industrial fermentation bio products (POCI-01-0247-FEDER-027578).ResultsUrinary tract infections (UTIs) are an increasingly common public health problem, with uropathogenic Escherichia coli (UPEC) being the most common etiological agent. Thestandard treatment for chronic UTIs implies long-term prophylactic use of antibiotics which promotes the development of antibiotic resistant bacteria. D-mannose andmannanoligosaccharides (MOS) are regarded as a natural alternative to the prophylactic use of antibiotics to avoid UTI development, and therefore reduce the need forantibiotics. This strategy can be used for the prevention of recurrent UTIs, thus opening a new market section for D-mannose/MOS as health supplements, the noveltyregarding the use of MOS. The objective of this study was to evaluate two different MOS extracts’ capacity to affect UPEC’s infection of bladder cells (HTB-9), mimicking theinitial infection (competition), an established infection (treatment) and prophylaxis scenarios (through quantifying the viable cells of UPEC).Figure 1 - Cytotoxic evaluation of HTB-9 after 24 h of contact with the samples atconcentrations ranging from 2.5 to 30.0 mg/mL. Results are expressed percentage (%)metabolic inhibition in relation to the positive control. Assay in accordance with ISO10993-5 (2009)Figure 2 - Percentage (%) of adhesion inhibition of UPEC to HTB-9 after 2 h ofsimultaneous incubation with MOS 1, MOS 2, D-mannose and methyl mannose.Different letters indicate statistically significant differences (p < 0.05) between thedifferent samples. Results are expressed in relation to the control.Figure 3 - Percentage (%) of adhesion inhibition of UPEC to HTB-9 after 1 h, 2 h and 3 h ofHTB-9 incubation with MOS 1, MOS 2, D-mannose and methyl mannose for 1 h, 2 h and 3 h.Different letters indicate statistically significant differences (p < 0.05) between the differentsamples. Results are expressed in relation to the control.Figure 4 - Percentage (%) of inhibition of infection after 2 and 24 h in contact with MOS 1, MOS2, D-mannose, and methyl mannose. Different letters indicate statistically significant differences(p < 0.05) between the different samples (A,B,C,D for 2 h and a,b,c,d for 24 h). Results areexpressed in relation to the control.➢ Competition Assay❑ Samples + HTB-9 + UPEC           2 h at 37 ºC with 5% CO2Human Bladdercarcinoma cell line(HTB-9)Escherichia coli strain CFT073(UPEC)➢ Treatment Assay❑ HTB-9 +UPEC           2 h at 37 ºCSamples addition 2 and 24 h at 37ºC with 5% CO2➢ Prophylaxis Assay❑ Samples + HTB-9 1, 2 and 3 h at 37 ºC with 5% of CO2UPEC addition 2 h at 37 ºC with 5% CO2Samples tested at 2.5 mg/mLof mannose content (noncytotoxic concentration)HTB-9 seeding UPEC inoculated at MOI 10Samples:MOS 1 and MOS 2MannoseMethly MannoseSerial dilutions plated (PCA) and incubate at 37 ºC for 24 h 

Effect of mannan oligosaccharides extracts in uropathogenic escherichia coli adhesion in human bladder cells

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Effect of mannan oligosaccharides extracts in uropathogenic escherichia coli adhesion in human bladder cells

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