Antimicrobial against tetracyclines and encoded by a variety of

Antimicrobial Effect

Since the inception of antibiotics by Sir
Alexander Fleming, who isolated Penicillium
notatum which successfully inhibited the growth of Staphylococcus aureus 1 in 1928. The gradual fruition of novel
antibiotics has been marred by the eventuality of microbial resistance, with the
subsequent identification of Penicillium Resistant Staphylococcus in 1940 2. The
extent of antimicrobial resistance (AMR) is swiftly becoming a pandemic with
the potential of affecting the entire human population. Furthermore, antibiotic
effect differs greatly depending on bacterial species. For example, the
majority of strains of Streptococcus
pneumoniae in Britain are inhibited by 0.01?mg/l of benzylpenicillin to
meet the minimum inhibitory concentration, whilst for Escherichia coli
32-64?mg/l are required to meet the minimum inhibitory concentration, a quantity
of benzylpenicillin that is unadvisable in humans 3. Pharmaceutical companies
and academia have reduced their research into novel antibiotics, reducing the
possibility of new discoveries 4,5. It has been predicted by the World Health
Organisation that if a new alternative to antibiotics is not found in the
foreseeable future 10 million people could die in 2050,
due to microbial resistance, with a person dying every three seconds 6. Consequently,
reverting society back to an era where AMR deaths devastated populations

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Antibiotic Resistance

The growing lethality and coupled genetic
plasticity of bacteria and pathogens is due to a multitude of factors;
horizontal transfer of antimicrobial resistance genes facilitates bacterial
evolution through conjugation, transduction and transformation. Alternatively,
AMR bacteria can develop mutational resistance which can be sorted into four
basic mechanisms, i) reduced amount
of antibiotic assimilation, ii) modification
of antimicrobial target, iii) antimicrobial
genes alter transcription and translation of relevant bacterial strains
resulting in changes in global metabolism and iv) efflux of antibiotics through transport pumps which is
particularly prevalent against tetracyclines and encoded by a variety of genes
such as tet(A) 3.






Antimicrobial Peptides

The identification of antimicrobial
peptides (AMPs), were extracted from the rumen microbiome 7 from microorganisms
that produce them as defence against foreign encroaching microbes 8. This could
fill the potential void of antimicrobials by combating the growing threat of
AMR bacteria. AMPs are composed of different amounts of peptides dependent on
the type. The polymeric nature of these oligopeptides is derived from the amino
acids which range from five to over a hundred. Apart from their bactericidal
properties AMPs influence the healing of vascular injuries, chemotaxis and the
formation of new blood vessels from pre-existing vessels 9. AMPs have a
positive charge allowing for their bypass into negatively charged bacterial
membranes 8.


Practical Introduction

In this practical we tested Escherichia coli and Staphylococcus epidermis with three AMPs and the antibiotic Ciprofloxacin. Each bacterial
species was pipetted on to three respective 96 well MIC plates containing an individual
Antimicrobial Peptide and the antibiotic Ciprofloxacin. Black paper was placed
under each plate to test for the turbidity of each individual well representing
bacterial growth. Conclusions could then be drawn once the correct MIC value
was obtained. Three AMPs were identified using BLASTp- Lynronne-1,
Lynronne-2 and Lynronne-3, Prevotella
ruminicola 23 (CP002006.1) is believed to be the producer of these AMPs.