Detection of diketopiperazine and pyrrolnitrin, compounds with anti-Pythium insidiosum activity, in a Pseudomonas stutzeri environmental strain

Abstract

Aims. Screening of bacterial flora for strains producing metabolites with inhibitory effects on the human pathogenic oomycete Pythium insidiosum. Separation and characterization of extracts from Pseudomonas stutzeri with anti-Pythium inhibitory activity. Search for genes with anti-Pythium effect within the genome of P. stutzeri. Methods. A total of 88 bacterial strains were isolated from water resources in northeastern Thailand. Two screening methods were used to establish their inhibitory effects on P. insidiosum. One strain, P. stutzeri ST1302 was randomly chosen, and the extract with anti-P. insidiosum activity was fractionated and subfractionated using liquid column chromatography and purified by thin layer chromatography. The chemical structure of purified fractions was determined by Fourier transform infrared spectroscopy, nuclear magnetic resonance and mass spectrometry. Further, search for genes involved in the anti-Pythium activity (phenazine-1-carboxylic acid, 2,4-diacetylphloroglucinol, pyoluteorin and pyrrolnitrin) was undertaken in this P. stutzeri strain using primers described in the literature. Results. Anti-P. insidiosum activity was detected in 16 isolates (18.2%). In P. stutzeri ST1302, a subfraction labeled PYK7 exhibited strong activity against this oomycete. It was assigned to the diketopiperazines as cyclo(D-Pro-L-Val). In the search for genes, one gene region was successfully amplified. This corresponded to pyrrolnitrin. The results suggest the possibility of using the related metabolites against P. insidiosum. This is the first report on the inhibitory effects of P. stutzeri against this oomycete. The results may contribute to the development of antimicrobial drugs/probiotics against pythiosis.

Evaluation of antifungal effect and toxicity of xanthyletin and two bacterial metabolites against Thai isolates of Pythium insidiosum

“…It is well known that the therapy of pythiosis has posed a challenge in recent decades as the current treatment options are highly variable and often lead to failure 14 . Therefore, many studies have searched for antimicrobial agents, natural compounds or secondary metabolites from microorganisms against P. insidiosum and results of in vitro testing are quite promising 6,7,[15][16][17][18][19][20][21] . The present study evaluated antimicrobial effects against P. insidiosum of xanthyletin and two fractions of the crude extracts from P. stutzeri ST1302 and K. pneumoniae ST2501.…”

“…Therefore, the treatment of pythiosis is still challenging. Previous studies demonstrated that natural compounds from plants and secondary metabolites from microorganisms had relatively good in vitro antimicrobial activity against P. insidiosum 6,7 . Xanthyletin, a natural compound of green plants, is classified in a group of pyranocoumarins, coumarin derivatives.…”

“…Moreover, microorganisms of the genus Klebsiella produce secondary metabolites with antibacterial and antifungal activity as well 11 . An in vitro study of secondary metabolites excreted from bacterial environmental strains included Pseudomonas stutzeri and Klebsiella pneumoniae from 16 strains that expressed anti-P. insidiosum activity 7 . However, the mechanisms of action of both xanthyletin and secondary metabolites from bacteria with anti-P. insidiosum activity remain unclear.…”

Identification and bioactive potential of marine microorganisms from selected Florida coastal areas

“…One such example includes the isolation of chemical entities from other P. stutzeri species that have yielded compounds such as alkyl-substituted diketopiperazines, pyrazines, and siderophores (Figure 4; Bañeras et al, 2013;Zawadzka et al, 2006). The diketoperazine that was isolated from the P. stutzeri strain ST1302 was shown to be active against the pathogenic fungus-like Pythium insidiosum using the disk diffusion method (Thongsri et al, 2014). The P. stutzeri strain (40M1) which was active in our bioassay against both the Gram-positive S. aureus and B. subtilis (Table 3) The different sample collection sites revealed minor variation in their microbial diversity.…”

A decade of understanding secondary metabolism in Pseudomonas spp. for sustainable agriculture and pharmaceutical applications

“…Many DKPs were considered as fermentation artifacts earlier, but now their antifungal, anti-mitotic and antibacterial roles have been unveiled. P. stutzeri showed the production of several DKPs with antiPythium insidiosum activity which is a mammalian pathogen (Thongsri et al 2014). DKPs mixture containing cyclo-lPro-Met, cyclo-l-Pro-l-Phe and cyclo-l-Pro-Val ( Fig.…”