Synergistic Killing of Pathogenic Escherichia coli Using Camel Lactoferrin from Different Saudi Camel Clans and Various Antibiotics

  • Hussein A. Almehdar
  • Nawal Abd El-Baky
  • Abdulqader A. Alhaider
  • Saud A. Almuhaideb
  • Abdullah A. Alhaider
  • Raed S. Albiheyri
  • Vladimir N. Uversky
  • Elrashdy M. RedwanEmail author


Current study aimed to analyze the synergistic killing of pathogenic Escherichia coli using camel lactoferrin from different Saudi camel clans and various antibiotics. Methods: using multiple microbiological and protein analysis techniques, the results were shown that the purified camel lactoferrins (cLfs) from different Saudi camel have strong antimicrobial potentials against two strains of E. coli. Although all cLfs were superior relative to human or bovine lactoferrins (hLf or bLf), there was no noticeable difference in the antimicrobial potentials of cLfs from different camel clans. The effects of antibiotics and cLfs were synergistic, indicating the superiority of using cLf-antibiotic combinations against E. coli growth. Since these combinations possessed distinguished synergy profiles, it is likely that they can be used to enhance the low efficacy of antibiotics, as well as to control the problems associated with bacterial resistance. Furthermore, these combinations can reduce the cost of cure of bacterial infections, especially in the developing countries. The analysis of the molecular mechanisms of lactoferrin action revealed that expression of several E. coli proteins was affected by the treatment with these antibacterial factors. Several proteins of different molecular weights interacting with cLf-biotin were found. Scanning and transmission electron microscopy analysis revealed the presence of noticeable morphological changes associated with the treatment of E. coli strains by antibiotic carbenicillin or cLf alone, and in combination. Camel lactoferrin has superior potential killing of E. coli over bovine and human lactoferrin, and this potential can be further synergistically enhanced of cLF is combined with antibiotics.


Camel clans Lactoferrins Antibiotics Synergy Escherichia coli Antimicrobial 


Author Contributions

EMR conceived the idea, supervised the project, organized and analyzed data, contributed to discussion, and with NAE-B wrote the manuscript and edited the manuscript. NAE-B did the experimental work, HAA, and AAA, VNU collected and analyzed data, contributed to discussion, and participated in writing and finalize the manuscript.


This work was supported by the King Abdulaziz City for Science and Technology General Directorate of Research Grants Programs, under Grant No. LGP-35-84.

Compliance with Ethical Standards

Competing Interests

The authors have no competing interests as defined by Nature Research, or other interests that might be perceived to influence the results and/or discussion reported in this paper.

Supplementary material

10930_2019_9828_MOESM1_ESM.docx (3.9 mb)
Supplementary material 1 (DOCX 3995 kb)


  1. 1.
    Black RE (1993) Epidemiology of diarrhoeal disease: implications for control by vaccines. Vaccine. 11(2):100–106Google Scholar
  2. 2.
    Scallan E, Mahon BE, Hoekstra RM, Griffin PM (2013) Estimates of illnesses, hospitalizations and deaths caused by major bacterial enteric pathogens in young children in the United States. Pediatr Infect Dis J. 32(3):217–221. Google Scholar
  3. 3.
    WHO (2006) Future directions for research on enterotoxigenic Escherichia coli vaccines for developing countries. Wkly Epidemiol Rec. 81(11):97–104Google Scholar
  4. 4.
    Walker CL, Black RE (2010) Diarrhoea morbidity and mortality in older children, adolescents, and adults. Epidemiol Infect 138(9):1215–1226. Google Scholar
  5. 5.
    Conway T, Cohen PS (2015) Commensal and pathogenic Escherichia coli metabolism in the gut. Microbiol Spectr. Google Scholar
  6. 6.
    Finegold SM, Sutter VL, Mathisen GE (1983) Normal indigenous intestinal microflora. In: Hentges DJ (ed) Human intestinal microflora in health and disease. Academic Press, Inc., New York, pp 3–31Google Scholar
  7. 7.
    Tenaillon O, Skurnik D, Picard B, Denamur E (2010) The population genetics of commensal Escherichia coli. Nat Rev Microbiol 8(3):207–217. Google Scholar
  8. 8.
    Wirth T, Falush D, Lan R, Colles F, Mensa P, Wieler LH et al (2006) Sex and virulence in Escherichia coli: an evolutionary perspective. Mol Microbiol 60(5):1136–1151. Google Scholar
  9. 9.
    Nataro JP, Kaper JB (1998) Diarrheagenic Escherichia coli. Clin Microbiol Rev 11(1):142–201Google Scholar
  10. 10.
    Redwan EM, Tabll A (2007) Camel lactoferrin markedly inhibits hepatitis C virus genotype 4 infection of human peripheral blood leukocytes. J Immunoassay Immunochem 28(3):267–277. Google Scholar
  11. 11.
    El Agamy EI (2006) Camel milk. In: Park YW, Haenlein FW (eds) Handbook of non-bovine mammals. Blackwell Publisher Professional, Iowa, pp 297–344Google Scholar
  12. 12.
    Farah Z, Mollet M, Younan M, Dahir R (2007) Camel dairy in Somalia: limiting factors and development potential. Livestock Sci. 110:187–191Google Scholar
  13. 13.
    Farah Z (1996) Camel milk properties and products. Swiss Centre for Developments Cooperation in Technology and Management, St. GallenGoogle Scholar
  14. 14.
    Zhang H, Yao J, Zhao D, Liu H, Li J, Guo M (2005) Changes in chemical composition of Alxa bactrian camel milk during lactation. J Dairy Sci 88(10):3402–3410. Google Scholar
  15. 15.
    Ramadan S, Inoue-Murayama M (2017) Advances in camel genomics and their applications: a review. J Anim Gen. 45:49–58Google Scholar
  16. 16.
    Kappeler S, Farah Z, Puhan Z (1998) Sequence analysis of Camelus dromedarius milk caseins. J Dairy Res 65(2):209–222Google Scholar
  17. 17.
    Konuspayeva G, Faye B, Loiseau G (2009) The composition of camel milk: a meta-analysis of the literature data. J Food Compost Anal. 22:95–101Google Scholar
  18. 18.
    Nikkah A (2011) Equidae, camel, and yak milks as functional foods: a review. J Nutr Food Sci. 1(5):1000116Google Scholar
  19. 19.
    Konuspayeva G, Faye B, Loiseau G, Levieux D (2007) Lactoferrin and immunoglobulin contents in camel’s milk (Camelus bactrianus, Camelus dromedarius, and Hybrids) from Kazakhstan. J Dairy Sci. 90(1):38–46Google Scholar
  20. 20.
    Pammi M, Abrams SA (2015) Oral lactoferrin for the prevention of sepsis and necrotizing enterocolitis in preterm infants. Cochrane Database Syst Rev. Google Scholar
  21. 21.
    Johnston WH, Ashley C, Yeiser M, Harris CL, Stolz SI, Wampler JL et al (2015) Growth and tolerance of formula with lactoferrin in infants through one year of age: double-blind, randomized, controlled trial. BMC Pediatr. 15:173. Google Scholar
  22. 22.
    Schlimme E, Martin D, Meisel H (2000) Nucleosides and nucleotides: natural bioactive substances in milk and colostrum. Br J Nutr 84(Suppl 1):S59–S68Google Scholar
  23. 23.
    Phelan M, Kerins D (2011) The potential role of milk-derived peptides in cardiovascular disease. Food Funct. 2(3–4):153–167. Google Scholar
  24. 24.
    Garcia C, Duan RD, Brevaut-Malaty V, Gire C, Millet V, Simeoni U et al (2013) Bioactive compounds in human milk and intestinal health and maturity in preterm newborn an overview. Cell Mol Biol. 59(1):108–131Google Scholar
  25. 25.
    Lonnerdal B (2014) Infant formula and infant nutrition: bioactive proteins of human milk and implications for composition of infant formulas. Am J Clin Nutr 99(3):712S–717S. Google Scholar
  26. 26.
    Hill DR, Newburg DS (2015) Clinical applications of bioactive milk components. Nutr Rev 73(7):463–476. Google Scholar
  27. 27.
    Hsieh CC, Hernandez-Ledesma B, Fernandez-Tome S, Weinborn V, Barile D, de Moura Bell JM (2015) Milk proteins, peptides, and oligosaccharides: effects against the 21st century disorders. Biomed Res Int. 2015:146840. Google Scholar
  28. 28.
    Sultan S, Huma N, Butt MS, Aleem M, Abbas M (2016) Therapeutic potential of dairy bioactive peptides: a contemporary perspective. Crit Rev Food Sci Nutr. 58(1):1–11Google Scholar
  29. 29.
    Demmelmair H, Prell C, Timby N, Lonnerdal B (2017) Benefits of lactoferrin, osteopontin and milk fat globule membranes for infants. Nutrients. 9(8):817Google Scholar
  30. 30.
    Marcone S, Belton O, Fitzgerald DJ (2017) Milk-derived bioactive peptides and their health promoting effects: a potential role in atherosclerosis. Br J Clin Pharmacol 83(1):152–162. Google Scholar
  31. 31.
    Conesa C, Calvo M, Sanchez L (2010) Recombinant human lactoferrin: a valuable protein for pharmaceutical products and functional foods. Biotechnol Adv 28(6):831–838. Google Scholar
  32. 32.
    Bruni N, Capucchio MT, Biasibetti E, Pessione E, Cirrincione S, Giraudo L et al (2016) Antimicrobial activity of lactoferrin-related peptides and applications in human and veterinary medicine. Molecules. 21(6):750Google Scholar
  33. 33.
    Jenssen H, Hamill P, Hancock RE (2006) Peptide antimicrobial agents. Clin Microbiol Rev 19(3):491–511. Google Scholar
  34. 34.
    Samyn-Petit B, Wajda Dubos JP, Chirat F, Coddeville B, Demaizieres G, Farrer S et al (2003) Comparative analysis of the site-specific N-glycosylation of human lactoferrin produced in maize and tobacco plants. Eur J Biochem. 270(15):3235–3242Google Scholar
  35. 35.
    El-Fakharany EM, Tabll A, Wahab AA, Redwan EM (2008) Potential activity of camel milk-amylase and lactoferrin against hepatitis C virus infectivity in HepG2 and lymphocytes. Hepat Mon. 8(2):101–109Google Scholar
  36. 36.
    El-Fakharany EM, Serour EA, Abdelrahman AM, Haroun BM, Redwan el RM (2009) Purification and characterization of camel (Camelus dromedarius) milk amylase. Prep Biochem Biotechnol 39(2):105–123. Google Scholar
  37. 37.
    Redwan EM (2009) Animal-derived pharmaceutical proteins. J Immunoassay Immunochem 30(3):262–290. Google Scholar
  38. 38.
    El-Fakharany EM, Haroun BM, Ng TB, Redwan ER (2010) Oyster mushroom laccase inhibits hepatitis C virus entry into peripheral blood cells and hepatoma cells. Protein Pept Lett 17(8):1031–1039Google Scholar
  39. 39.
    Almahdy O, El-Fakharany EM, El-Dabaa E, Ng TB, Redwan EM (2011) Examination of the activity of camel milk casein against hepatitis C virus (genotype-4a) and its apoptotic potential in hepatoma and hela cell lines. Hepat Mon. 11(9):724–730. Google Scholar
  40. 40.
    Ng TB, Wong JH, Almahdy O, El-Fakharany EM, El-Dabaa E, Redwan EM (2011) Antimicrobial activities of casein and other milk proteins. Casein: production, uses and health effects. Nova Publishers, New YorkGoogle Scholar
  41. 41.
    El-Fakharany EM, Abedelbaky N, Haroun BM, Sanchez L, Redwan NA, Redwan EM (2012) Anti-infectivity of camel polyclonal antibodies against hepatitis C virus in Huh7.5 hepatoma. Virol J. 9:201Google Scholar
  42. 42.
    Liao Y, El-Fakkarany E, Lonnerdal B, Redwan EM (2012) Inhibitory effects of native and recombinant full-length camel lactoferrin and its N and C lobes on hepatitis C virus infection of Huh7.5 cells. J Med Microbiol. 61(Pt 3):375–383Google Scholar
  43. 43.
    Conesa C, Sanchez L, Rota C, Perez MD, Calvo M, Farnaud S et al (2008) Isolation of lactoferrin from milk of different species: calorimetric and antimicrobial studies. Comp Biochem Physiol B 150(1):131–139Google Scholar
  44. 44.
    Redwan EM, El-Baky NA, Al-Hejin AM, Baeshen MN, Almehdar HA, Elsaway A et al (2016) Significant antibacterial activity and synergistic effects of camel lactoferrin with antibiotics against methicillin-resistant Staphylococcus aureus (MRSA). Res Microbiol 167(6):480–491. Google Scholar
  45. 45.
    Elass-Rochard E, Roseanu A, Legrand D, Trif M, Salmon V, Motas C et al (1995) Lactoferrin-lipopolysaccharide interaction: involvement of the 28-34 loop region of human lactoferrin in the high-affinity binding to Escherichia coli 055B5 lipopolysaccharide. Biochem J. 312(Pt 3):839–845Google Scholar
  46. 46.
    Almehdar HA, El-Baky NA, Alhaider AA, Almuhaideb SA, Alhaider AA, Albiheyri RS et al (2019) Bacteriostatic and bactericidal activities of camel lactoferrins against Salmonella enterica serovar typhi. Probiotics Antimicrob Proteins. Google Scholar
  47. 47.
    Albar AH, Almehdar HA, Uversky VN, Redwan EM (2014) Structural heterogeneity and multifunctionality of lactoferrin. Curr Protein Pept Sci 15(8):778–797Google Scholar
  48. 48.
    Wikler MA. Performance Standards for Antimicrobial Susceptibility Testing. Eighteenth Informational Supplement. Pennsylvania, PA, USA: C.L.S.I. (Clinical and Laboratory Standard Institute), 2008 Contract No.: M100-S18Google Scholar
  49. 49.
    Cockerill FR, Wikler MA, Alder J, Dudley MN, Eliopoulos GM, Ferraro MJ, et al. Methods for Dilution Antimicrobial Susceptibility Tests for Bacteria That Grow Aerobically; Approved Standard. CLSI document M07-A9.9. 9 ed. Pennsylvania, PA, USA: C.L.S.I. (Clinical and Laboratory Standard Institute); 2012Google Scholar
  50. 50.
    Redwan EM, Uversky VN, El-Fakharany EM, Al-Mehdar H (2014) Potential lactoferrin activity against pathogenic viruses. C R Biol. 337(10):581–595. Google Scholar
  51. 51.
    Park HM, Almeida RA, Oliver SP (2002) Identification of lactoferrin-binding proteins in Streptococcus dysgalactiae subsp. dysgalactiae and Streptococcus agalactiae isolated from cows with mastitis. FEMS Microbiol Lett. 207(1):87–90Google Scholar
  52. 52.
    Reddy SB, Mainwaring DE, Kobaisi MA, Zeephongsekul P, Fecondo JV (2012) Acoustic wave immunosensing of a meningococcal antigen using gold nanoparticle-enhanced mass sensitivity. Biosens Bioelectron 31(1):382–387. Google Scholar
  53. 53.
    Almeida RA, Luther DA, Park HM, Oliver SP (2006) Identification, isolation, and partial characterization of a novel Streptococcus uberis adhesion molecule (SUAM). Vet Microbiol 115(1–3):183–191. Google Scholar
  54. 54.
    Beddek AJ, Schryvers AB (2010) The lactoferrin receptor complex in Gram negative bacteria. Biometals 23(3):377–386. Google Scholar
  55. 55.
    Drago-Serrano ME, Campos-Rodriguez R, Carrero JC, de la Garza M (2017) Lactoferrin: balancing ups and downs of inflammation due to microbial infections. Int J Mol Sci. 18(3):501. Google Scholar
  56. 56.
    Drago-Serrano ME, de la Garza-Amaya M, Luna JS, Campos-Rodriguez R (2012) Lactoferrin-lipopolysaccharide (LPS) binding as key to antibacterial and antiendotoxic effects. Int Immunopharmacol 12(1):1–9. Google Scholar
  57. 57.
    Drago-Serrano ME, Rivera-Aguilar V, Resendiz-Albor AA, Campos-Rodriguez R (2010) Lactoferrin increases both resistance to Salmonella typhimurium infection and the production of antibodies in mice. Immunol Lett 134(1):35–46. Google Scholar
  58. 58.
    Morgenthau A, Partha SK, Adamiak P, Schryvers AB (2014) The specificity of protection against cationic antimicrobial peptides by lactoferrin binding protein B. Biometals 27(5):923–933. Google Scholar
  59. 59.
    Ochoa TJ, Cleary TG (2009) Effect of lactoferrin on enteric pathogens. Biochimie 91(1):30–34. Google Scholar
  60. 60.
    Rahman M, Kim WS, Kumura H, Shimazaki K (2009) Bovine lactoferrin region responsible for binding to bifidobacterial cell surface proteins. Biotechnol Lett 31(6):863–868. Google Scholar
  61. 61.
    Samaniego-Barron L, Luna-Castro S, Pina-Vazquez C, Suarez-Guemes F, de la Garza M (2016) Two outer membrane proteins are bovine lactoferrin-binding proteins in Mannheimia haemolytica A1. Vet Res 47(1):93. Google Scholar
  62. 62.
    Staggs TM, Greer MK, Baseman JB, Holt SC, Tryon VV (1994) Identification of lactoferrin-binding proteins from Treponema pallidum subspecies pallidum and Treponema denticola. Mol Microbiol. 12(4):613–619Google Scholar
  63. 63.
    Yu RH, Schryvers AB (2002) Bacterial lactoferrin receptors: insights from characterizing the Moraxella bovis receptors. Biochem Cell Biol. 80(1):81–90Google Scholar
  64. 64.
    Dhaenens L, Szczebara F, Husson MO (1997) Identification, characterization, and immunogenicity of the lactoferrin-binding protein from Helicobacter pylori. Infect Immun. 65(2):514–518Google Scholar
  65. 65.
    Fang W, Oliver SP (1999) Identification of lactoferrin-binding proteins in bovine mastitis-causing Streptococcus uberis. FEMS Microbiol Lett. 176(1):91–96Google Scholar
  66. 66.
    Schryvers AB, Morris LJ (1988) Identification and characterization of the human lactoferrin-binding protein from Neisseria meningitidis. Infect Immun. 56(5):1144–1149Google Scholar
  67. 67.
    Tomita S, Shirasaki N, Hayashizaki H, Matsuyama J, Benno Y, Kiyosawa I (1998) Binding characteristics of bovine lactoferrin to the cell surface of Clostridium species and identification of the lactoferrin-binding protein. Biosci Biotechnol Biochem 62(8):1476–1482. Google Scholar
  68. 68.
    Sharma AK, Paramasivam M, Srinivasan A, Yadav MP, Singh TP (1999) Three-dimensional structure of mare diferric lactoferrin at 2.6 A resolution. J Mol Biol. 289(2):303–317. Google Scholar
  69. 69.
    Sharma S, Sinha M, Kaushik S, Kaur P, Singh TP (2013) C-lobe of lactoferrin: the whole story of the half-molecule. Biochem Res Int. 2013:271641. Google Scholar
  70. 70.
    Sinha M, Kaushik S, Kaur P, Sharma S, Singh TP (2013) Antimicrobial lactoferrin peptides: the hidden players in the protective function of a multifunctional protein. Int J Pept. 2013:390230. Google Scholar
  71. 71.
    Khan JA, Kumar P, Paramasivam M, Yadav RS, Sahani MS, Sharma S et al (2001) Camel lactoferrin, a transferrin-cum-lactoferrin: crystal structure of camel apolactoferrin at 2.6 A resolution and structural basis of its dual role. J Mol Biol. 309(3):751–761Google Scholar
  72. 72.
    Khan JA, Kumar P, Srinivasan A, Singh TP (2001) Protein intermediate trapped by the simultaneous crystallization process. Crystal structure of an iron-saturated intermediate in the Fe3+ binding pathway of camel lactoferrin at 2.7 a resolution. J Biol Chem. 276(39):36817–36823. Google Scholar
  73. 73.
    Redwan EM, El-Fakharany EM, Uversky VN, Linjawi MH (2014) Screening the anti infectivity potentials of native N- and C-lobes derived from the camel lactoferrin against hepatitis C virus. BMC Complement Altern Med. 14:219. Google Scholar
  74. 74.
    Luo G, Spellberg B, Gebremariam T, Lee H, Xiong YQ, French SW et al (2014) Combination therapy with iron chelation and vancomycin in treating murine staphylococcemia. Eur J Clin Microbiol Infect Dis 33(5):845–851. Google Scholar

Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2019

Authors and Affiliations

  • Hussein A. Almehdar
    • 1
  • Nawal Abd El-Baky
    • 2
  • Abdulqader A. Alhaider
    • 1
  • Saud A. Almuhaideb
    • 3
  • Abdullah A. Alhaider
    • 4
  • Raed S. Albiheyri
    • 1
  • Vladimir N. Uversky
    • 1
    • 5
    • 6
  • Elrashdy M. Redwan
    • 1
    • 2
    Email author
  1. 1.Department of Biological Sciences, Faculty of SciencesKing Abdulaziz UniversityJeddahSaudi Arabia
  2. 2.Therapeutic and Protective Proteins Laboratory, Protein Research Department, Genetic Engineering and Biotechnology Research InstituteCity for Scientific Research and Technology ApplicationsAlexandriaEgypt
  3. 3.Tilad Veterinary CenterRiyadhSaudi Arabia
  4. 4.Medical SchoolKing Saud bin Abdulazziz University for Health ScienceRiyadhSaudi Arabia
  5. 5.Institute for Biological InstrumentationRussian Academy of Sciences of the PushchinoPushchinoRussia
  6. 6.Department of Molecular Medicine and USF Health Byrd Alzheimer’s Research Institute, Morsani College of MedicineUniversity of South FloridaTampaUSA

Personalised recommendations