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Taxonomic and Metabolomics Profiling of Actinobacteria Strains from Himalayan Collection Sites in Pakistan

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Abstract

Actinobacteria have proven themselves as the major producers of bioactive compounds with wide applications. In this study, 35 actinobacteria strains were isolated from soil samples collected from the Himalayan mountains region in Pakistan. The isolated strains were identified by polyphasic taxonomy and were prioritized based on biological and chemical screening to identify the strains with ability to produce inimitable metabolites. The biological screening included antimicrobial activity against Staphylococcus aureus, Micrococcus luteus, Salmonella enterica, Escherichia coli, Mycobacterium aurum, and Bacillus subtilis and anticancer activity using human cancer cell lines PC3 and A549. For chemical screening, methanolic extracts were investigated using TLC, HPLC-UV/MS. The actinobacteria strain PU-MM93 was selected for scale-up fermentation based on its unique chemical profile and cytotoxicity (50–60% growth inhibition) against PC3 and A549 cell lines. The scale-up fermentation of PU-MM93, followed by purification and structure elucidation of compounds revealed this strain as a promising producer of the cytotoxic anthracycline aranciamycin and aglycone SM-173-B along with the potent neuroprotective carboxamide oxachelin C. Other interesting metabolites produced include taurocholic acid as first report herein from microbial origin, pactamycate and cyclo(L-Pro-L-Leu). The study suggested exploring more bioactive microorganisms from the untapped Himalayan region in Pakistan, which can produce commercially significant compounds.

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References

  1. Takahashi Y, Nakashima T (2018) Actinomycetes, an inexhaustible source of naturally occurring antibiotics. Antibiotics 7:45. https://doi.org/10.3390/antibiotics7020045

    Article  CAS  PubMed Central  Google Scholar 

  2. Ibrahimi M, Korichi W, Hafidi M, Lemee L, Ouhdouch Y, Marine LS (2020) Actinobacteria: screening for predation leads to the discovery of potential new drugs against multidrug-resistant bacteria. Antibiotics 9:91. https://doi.org/10.3390/antibiotics9020091

    Article  PubMed Central  Google Scholar 

  3. Braesel J, Lee JH, Arnould B, Murphy BT et al (2019) Diazaquinomycin biosynthetic gene clusters from marine and freshwater actinomycetes. J Nat Prod 82:937–946. https://doi.org/10.1021/acs.jnatprod.8b01028

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  4. Fatima A, Aftab U, Shaaban KA, Thorson JS, Sajid I (2019) Spore forming actinobacterial diversity of Cholistan desert Pakistan: polyphasic taxonomy, antimicrobial potential and chemical profiling. BMC Microbiol 19(49):1–17. https://doi.org/10.1186/s12866-019-1414-x

    Article  Google Scholar 

  5. Goodfellow M, Nouioui I, Sanderson R, Xie F, Bull AT (2018) Rare taxa and dark microbial matter: novel bioactive actinobacteria abound in Atacama desert soils. Antonie Van Leeuwenhoek 111:1315–1332. https://doi.org/10.1007/s10482-018-1088-7

    Article  CAS  PubMed  Google Scholar 

  6. Mohammadipanah F, Wink J (2016) Actinobacteria from arid and desert habitats: diversity and biological activity. Front Microbiol 6:1541. https://doi.org/10.3389/fmicb.2015.01541

    Article  PubMed  PubMed Central  Google Scholar 

  7. Liu G, Chater KF, Chandra G, Niu G, Tan H (2013) Molecular regulation of antibiotic biosynthesis in Streptomyces. Microbiol Mol Biol Rev 77(1):112–143. https://doi.org/10.1128/MMBR.00054-12

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  8. Ibeyaima A, Rana J, Dwivedi AK, Saini N, Gupta S et al (2018) Pseudonocardiaceae sp. TD-015 from the Thar desert, India: antimicrobial activity and identification of antimicrobial compounds. Curr Bioact Compd 14:112–118. https://doi.org/10.2174/1573407213666170104124315

    Article  CAS  Google Scholar 

  9. Genilloud O (2017) Actinomycetes: still a source of novel antibiotics. Nat Prod Rep 34:1203–1232. https://doi.org/10.1039/C7NP00026J

    Article  CAS  PubMed  Google Scholar 

  10. Ziring L, Burki SJ (2020) Pakistan. In Encyclopedia Britannica 2020. https://www.britannica.com/place/Pakistan. Accessed 14 May 2020

  11. Shaheen H, Shinwari ZK, Qureshi RA, Ullah Z (2012) Indigenous plant resources and their utilization practices in village populations of Kashmir Himalayas. Pak J Bot 44:739–745

    Google Scholar 

  12. Ahmad KS, Habib S (2014) Indigenous knowledge of some medicinal plants of Himalaya region, Dawarian village, Neelum valley, Azad Jammu and Kashmir, Pakistan. Univ J Plant Sci 2:40–47. https://doi.org/10.13189/ujps.2014.020203

    Article  Google Scholar 

  13. Khan SM, Page S, Ahmad H, Shaheen H, Ullah Z, Ahmad M et al (2013) Medicinal flora and ethnoecological knowledge in the Naran valley, Western Himalaya, Pakistan. J Ethnobiol Ethnomed 9:4. https://doi.org/10.1186/1746-4269-9-4

    Article  PubMed  PubMed Central  Google Scholar 

  14. Abbas M, Elshahawi SI, Wang X, Ponomareva LV, Sajid I, Shaaban K et al (2018) Puromycins B-E, naturally occurring amino-nucleosides produced by the Himalayan isolate Streptomyces sp. PU-14G. J Nat Prod 81:2560–2566. https://doi.org/10.1021/acs.jnatprod.8b00720

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  15. Wang X, Abbas M, Zhang Y, Elshahawi SI, Ponomareva LV, Cui Z et al (2019) Baraphenazines A-G, divergent fused phenazine-based metabolites from a Himalayan Streptomyces. J Nat Prod 82:1686–1693. https://doi.org/10.1021/acs.jnatprod.9b00289

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  16. Hayakawa M, Yoshida Y, Iimura Y (2004) Selective isolation of bioactive soil actinomycetes belonging to the Streptomyces violaceusniger phenotypic cluster. J Appl Microbiol 96:973–981. https://doi.org/10.1111/j.1365-2672.2004.02230.x

    Article  CAS  PubMed  Google Scholar 

  17. Küster E, Williams S (1964) Selection of media for isolation of Streptomycetes. Nature 202:928–929. https://doi.org/10.1038/202928a0

    Article  Google Scholar 

  18. Shirling ET, Gottlieb D (1966) Methods for characterization of Streptomyces species. Int Bull Bact Nomen Toxonomy 16:313–340

    Google Scholar 

  19. Lane DJ, Stackebrandt E, Goodfellow M (1991) Nucleic acid techniques in bacterial systematics. Wiley, New York, pp 115–147

    Google Scholar 

  20. Wiegand I, Hilpert K, Hancock RE (2008) Agar and broth dilution methods to determine the minimal inhibitory concentration (MIC) of antimicrobial substances. Nat Protoc 3:163. https://doi.org/10.1038/nprot.2007.521

    Article  CAS  PubMed  Google Scholar 

  21. Wang X, Shaaban KA, Elshahawi SI, Ponomareva LV, Sunkara M, Zhang Y et al (2013) Frenolicins C-G, pyranonaphthoquinones from Streptomyces sp. RM-4-15. J Nat Prod 76:1441–1447. https://doi.org/10.1021/np400231r

    Article  CAS  PubMed  Google Scholar 

  22. Shaaban KA, Elshahawi SI, Wang X, Horn J, Kharel MK, Leggas M et al (2015) Cytotoxic indolocarbazoles from Actinomadura melliaura ATCC 39691. J Nat Prod 78:1723–1729. https://doi.org/10.1021/acs.jnatprod.5b00429

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  23. Voss SR, Ponomareva LV, Dwaraka VB, Pardue KE, Baddar NWAH, Rodgers AK et al (2019) HDAC regulates transcription at the outset of Axolotl tail regeneration. Sci Rep 9:1–11. https://doi.org/10.1038/s41598-019-43230-6

    Article  CAS  Google Scholar 

  24. Wang X, Elshahawi SI, Ponomareva LV, Ye Q, Liu Y, Copley GC et al (2019) Structure determination, functional characterization, and biosynthetic implications of nybomycin metabolites from a mining reclamation site-associated Streptomyces. J Nat Prod 12:3469–3476. https://doi.org/10.1021/acs.jnatprod.9b01015

    Article  CAS  Google Scholar 

  25. Zhang Y, Ye Q, Ponomareva LV, Cao Y, Liu Y, Cui Z et al (2019) Total synthesis of griseusins and elucidation of the griseusin mechanism of action. Chem Sci 10:7641–7648. https://doi.org/10.1039/C9SC02289A

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  26. Wang X, Zhang Y, Ponomareva LV, Qiu Q, Woodcock R, Elshahawi SI et al (2017) Mccrearamycins A-D, geldanamycin-derived cyclopentenone macrolactams from an Eastern Kentucky abandoned coal mine. Microbe Angew Chem Int Ed 129:3040–3044. https://doi.org/10.1002/ange.201612447

    Article  Google Scholar 

  27. Ponomareva LV, Athippozhy A, Thorson JS, Voss SR (2015) Using Ambystoma mexicanum (Mexican axolotl) embryos, chemical genetics, and microarray analysis to identify signaling pathways associated with tissue regeneration. Comp Biochem Physiol C 178:128–135. https://doi.org/10.1016/j.cbpc.2015.06.004

    Article  CAS  Google Scholar 

  28. Davies-Bolorunduro OF, Adeleye IA, Akinleye MO, Wang PG (2019) Anticancer potential of metabolic compounds from marine actinomycetes isolated from Lagos Lagoon sediment. J Pharm 9:201–208. https://doi.org/10.1016/j.jpha.2019.03.004

    Article  Google Scholar 

  29. Das R, Romi W, Das R, Sharma HK, Thakur D (2018) Antimicrobial potentiality of actinobacteria isolated from two microbiologically unexplored forest ecosystems of Northeast India. BMC Microbiol 18:71. https://doi.org/10.1186/s12866-018-1215-7

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  30. Bajaj BK, Sharma P (2011) An alkali-thermotolerant extracellular protease from a newly isolated Streptomyces sp. DP2. N Biotechnol 28:725–732. https://doi.org/10.1016/j.nbt.2011.01.001

    Article  CAS  PubMed  Google Scholar 

  31. Li J, Zhao GZ, Chen HH, Wang HB, Qin S, Zhu WY et al (2008) Antitumour and antimicrobial activities of endophytic streptomycetes from pharmaceutical plants in rainforest. Lett Appl Microbiol 47:574–580. https://doi.org/10.1111/j.1472-765X.2008.02470.x

    Article  CAS  PubMed  Google Scholar 

  32. Savi DC, Shaaban KA, Gos FM, Thorson JS, Glienke C, Rohr J (2019) Secondary metabolites produced by Microbacterium sp. LGMB471 with antifungal activity against the phytopathogen Phyllosticta citricarpa. Folia Microbiol 64:453–460. https://doi.org/10.1007/s12223-018-00668-x

    Article  CAS  Google Scholar 

  33. Elshahawi SI, Shaaban KA, Kharel MK, Thorson JS (2015) A comprehensive review of glycosylated bacterial natural products. Chem Soc Rev 44:7591–7697. https://doi.org/10.1039/C4CS00426D

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  34. Krohn K (2009) Anthracycline chemistry and biology I: biological occurence and biosynthesis, synthesis and chemistry. Springer, Heidelberg

    Google Scholar 

  35. Le Sann C (2006) Maleimide spacers as versatile linkers in the synthesis of bioconjugates of anthracyclines. Nat Prod Rep 23:357–367. https://doi.org/10.1039/B600666N

    Article  PubMed  Google Scholar 

  36. Keller-Schierlein W, Sauerbier J, Vogler U, Zähner H (1970) Metabolites of microorganisms. Aranciamycin Helv Chim Acta 53:779. https://doi.org/10.1002/hlca.19700530413

    Article  CAS  PubMed  Google Scholar 

  37. Bols M, Binderup L, Hansen J, Rasmussen P (1992) Inhibition of collagenase by aranciamycin and aranciamycin derivatives. J Med Chem 35:2768–2771. https://doi.org/10.1021/jm00093a008

    Article  CAS  PubMed  Google Scholar 

  38. Khalil ZG, Raju R, Piggott AM, Salim AA, Blumenthal A, Capon RJ (2015) Aranciamycins I and J, antimycobacterial anthracyclines from an Australian marine-derived Streptomyces sp. J Nat Prod 78:949–952. https://doi.org/10.1021/acs.jnatprod.5b00095

    Article  CAS  PubMed  Google Scholar 

  39. Fujiwara A, Hoshino T, Westley JW (1985) Anthracycline antibiotics. Crit Rev Biotechnol 3:133–157. https://doi.org/10.3109/07388558509150782

    Article  Google Scholar 

  40. Eida AA, Mahmud T (2019) The secondary metabolite pactamycin with potential for pharmaceutical applications: biosynthesis and regulation. Appl Microbiol 103:4337–4345. https://doi.org/10.1007/s00253-019-09831-x

    Article  CAS  Google Scholar 

  41. Shaaban KA, Saunders MA, Zhang Y, Tran T, Elshahawi SI, Ponomareva LV et al (2016) Spoxazomicin D and oxachelin C, potent neuroprotective carboxamides from the Appalachian coal fire-associated isolate Streptomyces sp. RM-14-6. J Nat Prod 80:2–11. https://doi.org/10.1021/acs.jnatprod.6b00948

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  42. Prasad C (1995) Bioactive cyclic dipeptides. Peptides 16:151–164. https://doi.org/10.1016/0196-9781(94)00017-Z

    Article  CAS  PubMed  Google Scholar 

  43. Zhai Y, Shao Z, Cai M, Zheng L, Li G, Yu Z et al (2019) Cyclo (l-Pro–l-Leu) of Pseudomonas putida MCCC 1A00316 isolated from Antarctic soil: identification and characterization of activity against Meloidogyne incognita. Molecules 24:768. https://doi.org/10.3390/molecules24040768

    Article  CAS  PubMed Central  Google Scholar 

  44. Rhee KH (2002) Isolation and characterization of Streptomyces sp. KH-614 producing anti-VRE (vancomycin-resistant enterococci) antibiotics. J Gen Appl Microbiol 48:321–327. https://doi.org/10.2323/jgam.48.321

    Article  CAS  PubMed  Google Scholar 

  45. Yan PS, Song Y, Sakuno E, Nakajima H, Nakagawa H, Yabe K (2004) Cyclo (L-leucyl-L-prolyl) produced by Achromobacter xylosoxidans inhibits aflatoxin production by Aspergillus parasiticus. Appl Environ Microbiol 70:7466–7473. https://doi.org/10.1128/AEM.70.12.7466-7473.2004

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  46. Degrassi G, Aguilar C, Bosco M, Zahariev S, Pongor S, Venturi V (2002) Plant growth-promoting Pseudomonas putida WCS358 produces and secretes four cyclic dipeptides: cross-talk with quorum sensing bacterial sensors. Curr Microbiol 45:250–254. https://doi.org/10.1007/s00284-002-3704-y

    Article  CAS  PubMed  Google Scholar 

  47. Hanf R (2019) Methods of treatment of cholestatic diseases. Google Patents

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Acknowledgements

This work was supported by Higher Education Commission (HEC) Pakistan grant (HEC-NRPU Project 2121). The work was also supported by National Institutes of Health grants R24 OD21479 (SRV, JST), R01 GM115261 (JST), the Center of Biomedical Research Excellence (COBRE) in Pharmaceutical Research and Innovation (CPRI, NIH P20 GM130456), the University of Kentucky College of Pharmacy, the University of Kentucky Markey Cancer Center and the National Center for Advancing Translational Sciences (Grant Nos. UL1TR000117, UL1TR001998). We thank the College of Pharmacy NMR Center (University of Kentucky) for NMR support.

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Authors and Affiliations

  1. Institute of Microbiology and Molecular Genetics, University of the Punjab, Lahore, Pakistan

    Mohsin T. Cheema & Imran Sajid

  2. Center for Pharmaceutical Research and Innovation, University of Kentucky, Lexington, KY, 40536, USA

    Mohsin T. Cheema, Larissa V. Ponomareva, Tao Liu, Jon S. Thorson & Khaled A. Shaaban

  3. Department of Pharmaceutical Sciences, College of Pharmacy, University of Kentucky, Lexington, KY, 40536, USA

    Mohsin T. Cheema, Larissa V. Ponomareva, Tao Liu, Jon S. Thorson & Khaled A. Shaaban

  4. Department of Natural Products Chemistry, School of Pharmacy, China Medical University, Shenyang, 110122, China

    Tao Liu

  5. Department of Neuroscience, University of Kentucky, Lexington, KY, 40536, USA

    S. Randal Voss

  6. Ambystoma Genetic Stock Center, University of Kentucky, Lexington, KY, 40536, USA

    S. Randal Voss

  7. Spinal Cord and Brain Injury Research Center, University of Kentucky, Lexington, KY, 40536, USA

    S. Randal Voss

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Contributions

MTC, LVP and KAS performed research and analyzed data; MTC and KAS wrote the first draft of the manuscript; TL and SRV contributed to experimental design and consultation; IS supervised the strains isolation and identification; JST, KAS and IS contributed to the experimental design, project oversight and manuscript preparation. All authors read and approved the manuscript.

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Correspondence to Khaled A. Shaaban or Imran Sajid.

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The authors declare the following competing financial interest: JST is a co-founder of Centrose (Madison, WI, USA).

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Cheema, M.T., Ponomareva, L.V., Liu, T. et al. Taxonomic and Metabolomics Profiling of Actinobacteria Strains from Himalayan Collection Sites in Pakistan. Curr Microbiol 78, 3044–3057 (2021). https://doi.org/10.1007/s00284-021-02557-y

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