Potential secondary metabolite analysis of soil Streptomyces sp. GMR22 and antibacterial assay on Porphyromonas gingivalis ATCC 33277

https://doi.org/10.19106/JMedSci005402202202

Hera Nirwati(1*), Ema Damayanti(2), Eti Nurwening Sholikhah(3), . Mustofa(4), Jaka Widada(5)

(1) Department of Microbiology, Faculty of Medicine, Public Health and Nursing, Universitas Gadjah Mada, Yogyakarta
(2) Research Center for Food Technology and Processing, National Research and Innovation Agency, Gunungkidul
(3) Department of Pharmacology and Therapy, Faculty of Medicine, Public Health and Nursing, Universitas Gadjah Mada, Yogyakarta
(4) Department of Pharmacology and Therapy, Faculty of Medicine, Public Health and Nursing, Universitas Gadjah Mada, Yogyakarta
(5) Department of Microbiology, Faculty of Agriculture, Universitas Gadjah Mada, Yogyakarta, Indonesia
(*) Corresponding Author

Abstract


Infectious diseases caused by oral pathogenic bacteria are currently a serious problem due to the increasing incidence of antimicrobial resistance. Streptomyces sp. GMR22, a soil actinobacterium which has large-genome size. In previous studies, it was known to have antifungal, and antibiofilm activity on Candida albicans. However, its antibacterial activity on oral pathogenic bacterium, Porphyromonas gingivalis is not clear. This study aimed to identify potential active compound based on genome mining analysis and to evaluate the antibacterial activity of GMR22 extract on P. gingivalis ATCC 33277. Potential active compounds and biosynthesis gene clusters were analysis using antiSMASH version 5. Antibacterial activity assay was carried out by the microdilution method on P. gingivalis ATCC 33277. Based on genome mining analysis polyketide synthase (PKS), the Streptomyces sp. GMR22 is the abundant BGCs (35%) and has large-predicted compounds which have antibiotic-antibacterial activity (22.9%). On antibacterial assay, chloroform extract of GMR22 at 7.8 – 62.5 µg/mL has high antibacterial activity on P. gingivalis compared to other extracts. Soil Streptomyces sp. GMR22 bacterium has biotechnological potential to produce active compounds for antibacterial.


Keywords


actinobacteria; antibacterial; Streptomyces; Porphyromonas gingivalis

Full Text:

PDF


References

Conrads G, Klomp T, Deng D, Wenzler JS, Braun A, Abdelbray MMH. The antimicrobial susceptibility of Porphyromonas gingivalis: genetic repertoire, global phenotype, and review of the literature. Antibiotics 2021; 10(12):1438. https://doi.org/10.3390/antibiotics10121438
2. Ardila CM, Bedoya-García JA. Antimicrobial resistance of Aggregatibacter actinomycetemcomitans, Porphyromonas gingivalis and Tannerella forsythia in periodontitis patients. J Glob Antimicrob Resist 2020; 22:215–18
https://doi.org/10.1016/j.jgar.2020.02.024
3. Larsen T. Susceptibility of Porphyromonas gingivalis in biofilms to amoxicillin, doxycycline and metronidazole. Oral Microbiol Immunol 2002; 17(5):267-71
https://doi.org/10.1034/j.1399-302x.2002.170501.x
4. Bérdy J. Thoughts and facts about antibiotics: Where we are now and where we are heading. Journal of Antibiotics 2012; 65(8):441
https://doi.org/10.1038/ja.2012.54
5. Undabarrena A, Ugalde JA, Seeger M, Cámara B. Genomic data mining of the marine actinobacteria Streptomyces sp. H-KF8 unveils insights into multi-stress related genes and metabolic pathways involved in antimicrobial synthesis. PeerJ 2017; 5:e2912
https://doi.org/10.7717/peerj.2912
6. Dalisay DS, Willams DE, Wang XL, Centko R, Chen J, Andersem JE. Marine sediment-derived Streptomyces bacteria from British Columbia, Canada are a promising microbiota resource for the discovery of antimicrobial natural products. PLoS One 2013; 8(10):77078
https://doi.org/10.1371/journal.pone.0077078
7. Nurjasmi R, Widada J, Ngadiman. Diversity of Actinomycetes at several forest types in Wanagama I Yogyakarta and their potency as a producer of antifungal compound. Indones. J Biotechnol 2009; 14(2):1196-205
https://doi.org/10.22146/ijbiotech.7813
8. Alimuddin, Widada J, Asmara W, Mustofa. Antifungal production of a strain of Actinomycetes spp isolated from the rhizosphere of Cajuput plant: selection and detection of exhibiting activity against tested fungi. Indones J Biotechnol 2011; 16(1):1-10
https://doi.org/10.22146/ijbiotech.7829
9. Ziemert N, Alanjary M, Weber T. The evolution of genome mining in microbes-a review. Natural Product Reports 2016; 33(8):988-1005
https://doi,org/10.1039/c6np00025h
10. Weber T, Charusanti P, Musiol-Kroll EM, Jiang X, Tong Y, Kim HU. Metabolic engineering of antibiotic factories: New tools for antibiotic production in actinomycetes. Trends Biotechnol 2015; 33(1):15-26
https://doi.org/ 10.1016/j.tibtech.2014.10.009
11. Ward AC, Allenby NE. Genome mining for the search and discovery of bioactive compounds: The Streptomyces paradigm. FEMS Microbiology Letters 2018; 365(24).
https://doi.org/10.1093/femsle/fny240
12. Kumar V, Bharti A, Gusain O, Bisht GS. Scanning electron microscopy of Streptomyces without use of any chemical fixatives. Scanning 2011; 33(6):446-9.
https://doi.org/10.1002/sca.20261
13. Blin K, Shaw S, Steinke K, Villebro R, Ziemert M, Lee SY, et al. AntiSMASH 5.0: Updates to the secondary metabolite genome mining pipeline. Nucleic Acids Res 2019; 47(W1):W81-7
https://doi.org/10.1093/nar/gkz310
14. Ghanem NB, Sabry SA, El-Sherif ZM, Abu El-Ela GA. Isolation and enumeration of marine actinomycetes from seawater and sediments in Alexandria. J Gen Appl Microbiol 2000; 46(3):105-11.
https://doi.org/10.2323/jgam.46.105
15. Carrol, D. H., Chassagne, F., Dettweiler, M. & Quave, C. L. Antibacterial activity of plant species used for oral health against Porphyromonas gingivalis. PLoS One 2020; 15(10):e0239316
https://doi.org/10.1371/journal.pone.0239316
16. Také A, Matsumoto A, Omura S, Takahashi Y. Streptomyces lactacystinicus sp. nov. and Streptomyces cyslabdanicus sp. nov., producing lactacystin and cyslabdan, respectively. J Antibiot (Tokyo) 2015; 68(11):719.
https://doi.org/10.1038/ja.2014.162
17. Herdini C, Mubarika S, Hariwiyanto B, Wijayanti N, Hosoyama A, Yamazoe A, et al.. Secondary bioactive metabolite gene clusters identification of anticandida-producing Streptomyces Sp. GMR22 isolated from Wanagama forest as revealed by Genome mining approach. Indones J Pharm 2017; 28(1):26-33.
http://dx.doi.org/10.14499/indonesianjpharm28iss1pp26
18. Baranasic D, Gacesa R, Starcevic A, Zucko J, Blazik M, Horvat M, et al. Draft genome sequence of Streptomyces rapamycinicus strain NRRL 5491, the producer of the immunosuppressant rapamycin. Genome Announc 2013; 1(4):e00581.
https://doi.org/10.1128/genomeA.00581-13
19. Wang XJ, Yan YJ, Zhang B, An J, Wang JJ, Tian J, et al. Genome sequence of the milbemycin-producing bacterium Streptomyces bingchenggensis. Journal of Bacteriology 2010; 192(17):4526-7.
https://doi.org/10.1128/JB.00596-10
20. Sun Y, Zhou X, Tu G, Deng Z. Identification of a gene cluster encoding meilingmycin biosynthesis among multiple polyketide synthase contigs isolated from Streptomyces nanchangensis NS3226. Arch Microbiol 180(2):101-7.
https://doi.org/10.1007/s00203-003-0564-1
21. Li L, Ma T, Liu Q, Huang Y, Hu C, Liao G. Improvement of daptomycin production in Streptomyces roseosporus through the acquisition of pleuromutilin resistance. Biomed Res Int 2013; 2013:479742.
https://doi.org/10.1155/2013/479742
22. Férir G, Hanchen A, Francois KO, Hoorelbeke B, Huskens D, Dettner F, et al. Feglymycin, a unique natural bacterial antibiotic peptide, inhibits HIV entry by targeting the viral envelope protein gp120. Virology 2012; 433(2):308-19.
https://doi.org/10.1016/j.virol.2012.08.007
23. Chandrakar S, Gupta AK. Actinomycin-producing endophytic Streptomyces parvulus associated with root of Aloe vera and optimization of conditions for antibiotic production. Probiotics Antimicrob Proteins 2019; 11(3)1055-69.
https://doi.org/10.1007/s12602-018-9451-6
24. Kong F, Carter GT. Structure determination of glycinosins A to D, further evidence for the cyclic structure of the amphomycin antibiotics. J Antibiot (Tokyo) 2003; 56(6):557-64.
https://doi.org/10.7164/antibiotics.56.557
25. Shacka JJ, Klocke BJ, Roth KA. Autophagy, bafilomycin and cell death: The ‘A-B-Cs’ of plecomacrolide-induced neuroprotection. Autophagy 2006; 2(3):228-30.
https://doi.org/10.4161/auto.2703
26. Jia XY, Tian ZH, Shao L, Qu XD, Zhao QF, Tang J, et al. Genetic Characterization of the Chlorothricin Gene Cluster as a Model for Spirotetronate Antibiotic Biosynthesis. Chem Biol 2006; 13(6):575-85.
https://doi.org/10.1016/j.chembiol.2006.03.008
27. Ichinose K, Ozawa M, Itou K, Kunieda K, Ebizuka Y. Cloning, sequencing and heterologous expression of the medermycin biosynthetic gene cluster of Streptomyces sp. AM-7161: Towards comparative analysis of the benzoisochromanequinone gene clusters. Microbiology 2003; 149(Pt 7):1633-45.
https://doi.org/10.1099/mic.0.26310-0
28. Ibrahim AA, El-Housseiny GS, Aboshanab KM, Yassien MA, Hassouna NA. Paromomycin production from Streptomyces rimosus NRRL 2455: Statistical optimization and new synergistic antibiotic combinations against multidrug resistant pathogens. BMC Microbiol 19; 19(1):18.
https://doi.org/10.1186/s12866-019-1390-1
29. Wu C, Shang Z, Lemetre C, Ternei MA, Brady SF. Cadasides, calcium-dependent acidic lipopeptides from the soil metagenome that are active against multidrug-resistant bacteria. J Am Chem Soc 2019; 141(9):3910-9.
https://doi.org/10.1021/jacs.8b12087
30. Yuan G, Xu L, Xu X, Li P, Zhong Q, Xia H, et al. Azalomycin F 5a , a polyhydroxy macrolide binding to the polar head of phospholipid and targeting to lipoteichoic acid to kill methicillin-resistant Staphylococcus aureus. Biomed Pharmacother 2019; 109:1940-50.
https://doi.org/10.1016/j.biopha.2018.11.067
31. McAlpine JB, Bachmann BO, Piraee M, Tremblay S, Alarco AM, Zazopoulos E, et al. Microbial genomics as a guide to drug discovery and structural elucidation: ECO-02301, a novel antifungal agent, as an example. J Nat Prod 2005; 68(4):493-6.
https://doi.org/10.1021/np0401664
32. Van Arnam EB, Ruzzini AC, Sit CS, Horn H, Tomas AAP, Currie CR, et al. Selvamicin, an atypical antifungal polyene from two alternative genomic contexts. Proc Natl Acad. Sci USA 2016; 113(46):12940-5.
https://doi.org/10.1073/pnas.1613285113
33. Sun F, Xu S, Jiang F, Liu W. Genomic-driven discovery of an amidinohydrolase involved in the biosynthesis of mediomycin A. Appl Microbiol Biotechnol 2018; 102(5):2225-34.
https://doi.org/10.1007/s00253-017-8729-z
34. Elsayed EA, Farid MA, El-Enshasy HA. Enhanced Natamycin production by Streptomyces natalensis in shake-flasks and stirred tank bioreactor under batch and fed-batch conditions. BMC Biotechnol 2019; 19(1):46.
https://doi.org/10.1186/s12896-019-0546-2
35. Alimuddin, Asmara W, Widada J, Nurjasmi R. An Actinomycetes Producing Anticandida Isolated from Cajuput Rhizosphere: Partial Identification of Isolates and Amplification of pks-I genes. Indones J Biotechnol 2010; 15(1):1-8.
https://doi.org/10.22146/ijbiotech.7817
36. Kumar R, Musiyenko A, Barik S. The heat shock protein 90 of Plasmodium falciparum and antimalarial activity of its inhibitor, geldanamycin. Malar J 2003; 2(30).
https://doi.org/10.1186/1475-2875-2-30
37. Neznanov N, Gorbachev AV, Neznanova L, Komarov AP, Gurova KV, Gasparian AV, et al. Anti-malaria drug blocks proteotoxic stress response: Anti-cancer implications. Cell Cycle 2009; 8(23):3960-70.
https://doi.org/10.4161/cc.8.23.10179
38. D’Alessandro S, Corbett Y, Ilboudo DP, Misiano P, Dahiya N, Abay SM, et al. Salinomycin and other ionophores as a new class of antimalarial drugs with transmission-blocking activity. Antimicrob Agents Chemother 2015; 59(9):5135-44.
https://doi.org/10.1128/AAC.04332-14



DOI: https://doi.org/10.19106/JMedSci005402202202

Article Metrics

Abstract views : 1404 | views : 1168




Copyright (c) 2022 Hera Nirwati, Ema Damayanti, Eti Nurwening Sholikhah, . Mustofa, Jaka Widada

Creative Commons License
This work is licensed under a Creative Commons Attribution-NonCommercial 4.0 International License.

View My Stats

 

Creative Commons License
Journal of the Medical Sciences (Berkala Ilmu Kedokteran) by  Universitas Gadjah Mada is licensed under a Creative Commons Attribution-NonCommercial 4.0 International License.
Based on a work at http://jurnal.ugm.ac.id/bik/.

 

https://bkim.jatengprov.go.id/wp-content/product/bonus100/ https://bkim.jatengprov.go.id/wp-content/product/max/ https://conference.ut.ac.id/pulsa/ https://conference.ut.ac.id/ovo/ http://jif.fmipa.unand.ac.id/docs/xbolax/ http://jif.fmipa.unand.ac.id/docs/sv388/ http://jmua.fmipa.unand.ac.id/product/20-thai/ http://jmua.fmipa.unand.ac.id/product/bonus120/ http://jmua.fmipa.unand.ac.id/product/totoslt/ http://jpt.faperta.unand.ac.id/file/xpulsa/ http://jpt.faperta.unand.ac.id/file/xtogel/ http://jurnaldampak.ft.unand.ac.id/docs/sv388x/ https://bkim.jatengprov.go.id/wp-content/uploads/thaix/ http://ocs.wima.ac.id/pages/xdana/ http://ocs.wima.ac.id/pages/xqris/ http://ocs.ukwms.ac.id/public/xbandar/ http://ocs.ukwms.ac.id/public/xbola/ http://ocs.ukwms.ac.id/public/xtoto/ http://jurnaldampak.ft.unand.ac.id/docs/toto-slt/ http://jurnaldampak.ft.unand.ac.id/docs/xdemo/ http://jurnal.wima.ac.id/pages/bonus100/ http://jurnal.wima.ac.id/pages/qris/ https://simkatmawa.untan.ac.id/wp-content/themes/sunda/ http://ruangjuru.dpuair.jatimprov.go.id/public/file/bonusx/ http://ruangjuru.dpuair.jatimprov.go.id/public/file/pulsax/ http://ruangjuru.dpuair.jatimprov.go.id/public/file/xthaix/ https://jurnal.fkip.unila.ac.id/pages/demo/ https://jurnal.fkip.unila.ac.id/pages/thai/ https://library.iptrisakti.ac.id/uploads/thai/ https://library.iptrisakti.ac.id/uploads/bonus/ https://library.iptrisakti.ac.id/uploads/max/ https://library.iptrisakti.ac.id/uploads/pulsa/ https://puskap.iptrisakti.ac.id/wp-content/upload/kambo/ https://paramadina.ac.id/media/qris/ https://paramadina.ac.id/media/ovo/ https://paramadina.ac.id/media/gopay/ https://jurnal.fkip.unila.ac.id/js/kambox/ https://puskap.iptrisakti.ac.id/wp-content/pulsax/ https://puskap.iptrisakti.ac.id/wp-content/bonusx/ https://ruangjuru.dpuair.jatimprov.go.id/public/assets/xkambox/ https://ruangjuru.dpuair.jatimprov.go.id/public/assets/xmax/ https://eoffice.uki.ac.id/upload/users/000/thai/ https://jurnal.pascasarjana.uniba-bpn.ac.id/files/xqris/ https://jurnal.pascasarjana.uniba-bpn.ac.id/files/xpulsa/ https://jurnal.pascasarjana.uniba-bpn.ac.id/files/xbonus/ https://icoresct.uniba-bpn.ac.id/pages/totox/ https://icoresct.uniba-bpn.ac.id/pages/ogelx/ https://icoresct.uniba-bpn.ac.id/pages/bdslt/ http://jbma.fk.unand.ac.id/js/okambo/ http://jbma.fk.unand.ac.id/js/otg/ http://jbma.fk.unand.ac.id/js/obd/ http://jbma.fk.unand.ac.id/js/ototo/ http://jfu.fmipa.unand.ac.id/store/dana/ http://jfu.fmipa.unand.ac.id/store/gopay/ http://jfu.fmipa.unand.ac.id/store/ovo/ http://jfu.fmipa.unand.ac.id/store/qris/ http://jif.fmipa.unand.ac.id/product/bonus/ http://jif.fmipa.unand.ac.id/product/demo/ http://jif.fmipa.unand.ac.id/product/kambo/ http://jif.fmipa.unand.ac.id/product/thai/ http://jrs.ft.unand.ac.id/icon/xdemox/ http://jrs.ft.unand.ac.id/icon/xbslotx/ http://jrs.ft.unand.ac.id/icon/xtogx/ http://jurnal.fk.unand.ac.id/event/zkambo/ http://jurnal.fk.unand.ac.id/event/zbonus/ http://jurnal.fk.unand.ac.id/event/zmax/ http://jurnal.fk.unand.ac.id/event/zthai/ http://jinh.fmipa.unand.ac.id/lib/malaysia/ http://jinh.fmipa.unand.ac.id/lib/singapore/ http://jinh.fmipa.unand.ac.id/lib/myanmar/ http://jinh.fmipa.unand.ac.id/lib/sv388/ http://jpt.faperta.unand.ac.id/docs/777/ http://jpt.faperta.unand.ac.id/docs/mahjongx/ http://jpt.faperta.unand.ac.id/docs/pg/ http://jpt.faperta.unand.ac.id/docs/maxwin/ https://jurnal.fkip.unila.ac.id/store/zthai/ https://jurnal.fkip.unila.ac.id/store/ztoto/ https://jurnal.fkip.unila.ac.id/store/zmax/ https://jurnal.fkip.unila.ac.id/store/zbonus/ http://jurnalmka.fk.unand.ac.id/menu/xqrisx/ http://jurnalmka.fk.unand.ac.id/menu/xbdblx/ http://jurnalmka.fk.unand.ac.id/menu/xbdtgx/ http://jurnalmka.fk.unand.ac.id/menu/xbdsltx/ https://bkim.jatengprov.go.id/wp-admin/css/totox/ https://bkim.jatengprov.go.id/wp-admin/css/demox/ https://jurnal.fkip.unila.ac.id/qrisx/ https://jurnal.fkip.unila.ac.id/bonusx/ https://jurnal.fkip.unila.ac.id/pulsax/ https://logistik.unsia.ac.id/xbonus/ https://logistik.unsia.ac.id/xgacor/ https://logistik.unsia.ac.id/xthai/ https://logistik.unsia.ac.id/xpulsa/ http://alumni.uinjambi.ac.id/uploadss/totosl/ http://alumni.uinjambi.ac.id/uploadss/bdsl/ http://alumni.uinjambi.ac.id/uploadss/bdtgl/ http://alumni.uinjambi.ac.id/uploadss/sgacor/ http://portalsiakad.uinjambi.ac.id/uploadss/demo/ http://portalsiakad.uinjambi.ac.id/uploadss/thai/ http://portalsiakad.uinjambi.ac.id/uploadss/mahjong/ http://portalsiakad.uinjambi.ac.id/uploadss/resmi/ http://pembayaran.uinjambi.ac.id/s/thai/ http://pembayaran.uinjambi.ac.id/s/bonus/ http://pembayaran.uinjambi.ac.id/s/pulsa/ http://registrasi.uinjambi.ac.id/uploadss/xbtogel/ http://registrasi.uinjambi.ac.id/uploadss/xbslot/ http://registrasi.uinjambi.ac.id/uploadss/xbtoto/ http://siakad.uinjambi.ac.id/uploadss/demo/ http://siakad.uinjambi.ac.id/uploadss/max/ http://siakad.uinjambi.ac.id/uploadss/kambo/ https://api.register.ppid.sijunjung.go.id/-/kamboja/ https://api.register.ppid.sijunjung.go.id/-/thai/ https://api.register.ppid.sijunjung.go.id/-/bonus/ https://api.register.ppid.sijunjung.go.id/-/maxwin/ https://bappeda.sijunjung.go.id/wp-content/product/totox/ https://bappeda.sijunjung.go.id/wp-content/product/bdtog/ https://bappeda.sijunjung.go.id/wp-content/product/scatter/ https://bappeda.sijunjung.go.id/wp-content/product/mahjong/ https://baznas.sijunjung.go.id/wp-content/uploads/demo/ https://baznas.sijunjung.go.id/wp-content/uploads/kambo/ https://baznas.sijunjung.go.id/wp-content/uploads/max/ https://baznas.sijunjung.go.id/wp-content/uploads/myanmar/ https://bbmsetda.sijunjung.go.id/thai/ https://bbmsetda.sijunjung.go.id/777/ https://bbmsetda.sijunjung.go.id/xbonusx/ https://bbmsetda.sijunjung.go.id/bandars/ https://bkpsdm.sijunjung.go.id/wp-content/xsingaporex/ https://bkpsdm.sijunjung.go.id/wp-content/xsateslotx/ https://bkpsdm.sijunjung.go.id/wp-content/xladangx/ https://bkpsdm.sijunjung.go.id/wp-content/xvipx/ https://bpbd.sijunjung.go.id/wp-content/xbbolax/ https://bpbd.sijunjung.go.id/wp-content/xtotox/ https://bpbd.sijunjung.go.id/wp-content/xbtogelx/ https://bpbd.sijunjung.go.id/wp-content/xsv388x/ https://rsud.sijunjung.go.id/wp-content/xbdx/ https://rsud.sijunjung.go.id/wp-content/xdemox/ https://rsud.sijunjung.go.id/wp-content/xkambox/ https://rsud.sijunjung.go.id/wp-content/xmax/ https://nagarimuarotakung.sijunjung.go.id/wp-content/shop/zbonus/ https://nagarimuarotakung.sijunjung.go.id/wp-content/shop/zkambo/ https://nagarimuarotakung.sijunjung.go.id/wp-content/shop/zmax/ https://nagarimuarotakung.sijunjung.go.id/wp-content/shop/zpulsa/ https://dekranasda.sijunjung.go.id/wp-content/bonus100/ https://dekranasda.sijunjung.go.id/wp-content/pulsa100/ https://dekranasda.sijunjung.go.id/wp-content/maxwin/ https://dekranasda.sijunjung.go.id/wp-content/thailand/ https://dinkes.sijunjung.go.id/wp-content/uploads/xbolax/ https://dinkes.sijunjung.go.id/wp-content/uploads/xgacorx/ https://dinkes.sijunjung.go.id/wp-content/uploads/xscatterx/ https://dinkes.sijunjung.go.id/wp-content/uploads/xzeusx/ https://disparpora.sijunjung.go.id/wp-content/uploads/oakuno/ https://disparpora.sijunjung.go.id/wp-content/uploads/obonuso/ https://disparpora.sijunjung.go.id/wp-content/uploads/odemo/ https://disparpora.sijunjung.go.id/wp-content/uploads/omaxo/ https://geoparksilokek.sijunjung.go.id/wp-content/uploads/bandark/ https://geoparksilokek.sijunjung.go.id/wp-content/uploads/kambo/ https://geoparksilokek.sijunjung.go.id/wp-content/uploads/pulsak/ https://geoparksilokek.sijunjung.go.id/wp-content/uploads/totok/ https://jpipm.uho.ac.id/public/bonust/ https://jpipm.uho.ac.id/public/pulsat/ https://jpipm.uho.ac.id/public/maxt/ https://jpipm.uho.ac.id/public/thait/ https://jpfi.uho.ac.id/docs/bandarsk/ https://jpfi.uho.ac.id/docs/bandartk/ https://jpfi.uho.ac.id/docs/kambok/ https://jpfi.uho.ac.id/docs/totok/ https://agroteknos.uho.ac.id/lib/demok/ https://agroteknos.uho.ac.id/lib/maxwin/ https://agroteknos.uho.ac.id/lib/scatter/ https://agroteknos.uho.ac.id/lib/zeusx/ https://celebica.uho.ac.id/pages/bola/ https://celebica.uho.ac.id/pages/mahjong/ https://celebica.uho.ac.id/pages/akun/ https://celebica.uho.ac.id/pages/sv388/ https://ecoforest.uho.ac.id/docs/xbonusx/ https://ecoforest.uho.ac.id/docs/xpulsax/ https://ecoforest.uho.ac.id/docs/xqrisx/ https://ecoforest.uho.ac.id/docs/xthaix/ https://edusains.uho.ac.id/icon/777/ https://edusains.uho.ac.id/icon/demox/ https://edusains.uho.ac.id/icon/gacor4d/ https://edusains.uho.ac.id/icon/gacorx/ https://jurnal.pbsi.uniba-bpn.ac.id/public/xtotox/ https://jurnal.pbsi.uniba-bpn.ac.id/public/xbolax/ https://jurnal.pbsi.uniba-bpn.ac.id/public/xqrisx/ https://jurnal.peko.uniba-bpn.ac.id/docs/xthaix/ https://jurnal.peko.uniba-bpn.ac.id/docs/xkambox/ https://jurnal.peko.uniba-bpn.ac.id/docs/xmalayx/ https://jurnal.pmat.uniba-bpn.ac.id/pages/xovox/ https://jurnal.pmat.uniba-bpn.ac.id/pages/xmahjongx/ https://jurnal.pmat.uniba-bpn.ac.id/pages/xgopayx/ https://kompetensi.fkip.uniba-bpn.ac.id/pages/bd-tg/ https://kompetensi.fkip.uniba-bpn.ac.id/pages/bd-slt/ https://kompetensi.fkip.uniba-bpn.ac.id/pages/777/ https://konteks17.uniba-bpn.ac.id/docs/xdemox/ https://konteks17.uniba-bpn.ac.id/docs/xbonusx/ https://konteks17.uniba-bpn.ac.id/docs/xpulsax/ http://kepaniteraan.fk.hangtuah.ac.id/wp-content/uploads/max/ http://kepaniteraan.fk.hangtuah.ac.id/wp-content/uploads/toto/ http://kepaniteraan.fk.hangtuah.ac.id/wp-content/uploads/bonusx/ https://sikd.untirta.ac.id/vendor/xdemox/ https://sikd.untirta.ac.id/vendor/xkambox/ https://sikd.untirta.ac.id/vendor/xthaix/ https://sikd.untirta.ac.id/vendor/xtotox/ https://irecoms.ump.ac.id/asset/zbonus/ https://irecoms.ump.ac.id/asset/zpulsa/ https://irecoms.ump.ac.id/asset/ztotox/ https://irecoms.ump.ac.id/asset/zbslot/ https://jurnal.unhamzah.ac.id/docs/pulsa/ https://jurnal.unhamzah.ac.id/docs/max/ https://jurnal.unhamzah.ac.id/docs/bonus/ https://jurnal.unhamzah.ac.id/docs/mahjong/ https://sistem.poltekindonusa.ac.id/wp-content/xbonusx/ https://sistem.poltekindonusa.ac.id/wp-content/xdemox/ https://sistem.poltekindonusa.ac.id/wp-content/xmax/ https://sistem.poltekindonusa.ac.id/wp-content/xpulsax/ https://diskominfo.sijunjung.go.id/wp-content/uploads/akun/ https://diskominfo.sijunjung.go.id/wp-content/uploads/mahjong2/ https://diskominfo.sijunjung.go.id/wp-content/uploads/resmi/ https://diskominfo.sijunjung.go.id/wp-content/uploads/server/ https://disnakertrans.sijunjung.go.id/wp-content/uploads/gacor4d/ https://disnakertrans.sijunjung.go.id/wp-content/uploads/malaysia/ https://disnakertrans.sijunjung.go.id/wp-content/uploads/pg/ https://disnakertrans.sijunjung.go.id/wp-content/uploads/sbobet/ https://inspektorat.sijunjung.go.id/wp-content/uploads/xbandarx/ https://inspektorat.sijunjung.go.id/wp-content/uploads/xbonusx/ https://inspektorat.sijunjung.go.id/wp-content/uploads/xpulsax/ https://inspektorat.sijunjung.go.id/wp-content/uploads/xmahjongx/ https://jurnal.fea.uniba-bpn.ac.id/pages/xmax/ https://jurnal.fea.uniba-bpn.ac.id/pages/xbonusx/ https://jurnal.fea.uniba-bpn.ac.id/pages/xqrisx/ https://jurnal.fem.uniba-bpn.ac.id/docs/xbolax/ https://jurnal.fem.uniba-bpn.ac.id/docs/xpulsax/ https://jurnal.fem.uniba-bpn.ac.id/docs/xtotox/ https://jurnal.fte.uniba-bpn.ac.id/doc/bd-tg/ https://jurnal.fte.uniba-bpn.ac.id/doc/bd-slt/ https://jurnal.fte.uniba-bpn.ac.id/doc/xovox/ https://jurnal.ftm.uniba-bpn.ac.id/pages/odemo/ https://jurnal.ftm.uniba-bpn.ac.id/pages/okambo/ https://jurnal.ftm.uniba-bpn.ac.id/pages/ogopay/ https://jurnal.industri.uniba-bpn.ac.id/docs/xzeus/ https://jurnal.industri.uniba-bpn.ac.id/docs/xgatot/ https://jurnal.industri.uniba-bpn.ac.id/docs/xmahjong/ https://jurnal.lp2m.uniba-bpn.ac.id/public/xthaix/ https://jurnal.lp2m.uniba-bpn.ac.id/public/xgaransix/ https://jurnal.lp2m.uniba-bpn.ac.id/public/xmyanmarx/ https://jak.uho.ac.id/docs/zbdsltz/ https://jak.uho.ac.id/docs/zbdtogz/ https://jak.uho.ac.id/docs/zmahjongz/ https://jak.uho.ac.id/docs/ztotoz/ https://jipfi.uho.ac.id/shop/kambo/ https://jipfi.uho.ac.id/shop/malaysia/ https://jipfi.uho.ac.id/shop/thai/ https://jipfi.uho.ac.id/shop/zeus/ https://jipsd.uho.ac.id/public/demox/ https://jipsd.uho.ac.id/public/bonusx/ https://jipsd.uho.ac.id/public/max/ https://jipsd.uho.ac.id/public/pulsax/ https://jkl-fkm.uho.ac.id/icon/bolax/ https://jkl-fkm.uho.ac.id/icon/mahjong/ https://jkl-fkm.uho.ac.id/icon/rtp/ https://jkl-fkm.uho.ac.id/icon/scatterx/ https://job.uho.ac.id/pages/polax/ https://job.uho.ac.id/pages/bandarx/ https://job.uho.ac.id/pages/gacor4d/ https://job.uho.ac.id/pages/maxwinx/ https://jolimpic.uho.ac.id/js/kbonus/ https://jolimpic.uho.ac.id/js/kdemo/ https://jolimpic.uho.ac.id/js/kpulsa/ https://jolimpic.uho.ac.id/js/kqris/ https://jristam.uho.ac.id/data/macau/ https://jristam.uho.ac.id/data/datahk/ https://jristam.uho.ac.id/data/paitohk/ https://jristam.uho.ac.id/data/datasgp/ https://jurnal-gikes.uho.ac.id/lib/gopayx/ https://jurnal-gikes.uho.ac.id/lib/idnslot/ https://jurnal-gikes.uho.ac.id/lib/mposlot/ https://jurnal-gikes.uho.ac.id/lib/ovox/ https://jurnal-wins.uho.ac.id/public/pg/ https://jurnal-wins.uho.ac.id/public/pulsax/ https://jurnal-wins.uho.ac.id/public/totox/ https://jurnal-wins.uho.ac.id/public/bandarx/ https://ejournalagribisnis.uho.ac.id/js/pay4d/ https://ejournalagribisnis.uho.ac.id/js/pg/ https://ejournalagribisnis.uho.ac.id/js/resmi/ https://ejournalagribisnis.uho.ac.id/js/rtp/ https://rb.kemenpora.go.id/wp-content/files/bonus/ https://rb.kemenpora.go.id/wp-content/files/maxwin/ https://rb.kemenpora.go.id/wp-content/files/thai/ https://rb.kemenpora.go.id/wp-content/files/gacor/