Investigation into Phage/Antibiotics Synergistic Effects on Infection

This report investigates the synergistic effects of combining phage therapy with antibiotics to combat infections, particularly those resistant to multiple drugs. It examines the notion of phage-antibiotic synergy (PAS), where sublethal concentrations of antibiotics can stimulate host bacteria in the virulent phase, enhancing phage activity. The report discusses the impact of PAS, including its effectiveness against Gram-positive bacteria like MRSA, and explores various mechanisms of action, such as cell elongation and increased antibiotic susceptibility. It also highlights strategies to promote PAS and addresses the rationale for researching this approach, emphasizing its potential to reduce antibiotic usage and combat antimicrobial resistance. The study also touches upon the adverse effects of synergistic drugs and concludes with recommendations for future research and application.

Pharmacotherapy Bacteriophage
therapeutics

Table of Contents
TITLE..............................................................................................................................................1
CHAPTER 1: INTRODUCTION (1500)........................................................................................1
Aim.........................................................................................................................................1
Objective.................................................................................................................................2
RATIONALE FOR THE RESEARCH...........................................................................................2
CHAPTER 2: LITERATURE REVIEW (3500).............................................................................2
Notion of synergistic effect of phage/antibiotics to combat infection...................................2
Impact of synergistic effect of phage/antibiotics to combat infection...................................5
Ways and strategies that promote synergistic effect of phage/antibiotics to combat infection8
CHAPTER 3: RESEARCH METHODOLOGY (2500)................................................................10
CHAPTER 4: RESULT AND DISCUSSION (1500)..................................................................15
CHAPTER 5: CONCLUSION AND RECOMMENDATION (1000)..........................................18
Conclusion............................................................................................................................18
Recommendation..................................................................................................................19
REFERENCES..............................................................................................................................22

TITLE
“Investigation into the synergistic effects of phage/antibiotics to combat infection”
CHAPTER 1: INTRODUCTION (1500)
The phage-antibiotic synergy is referring as process where sub lethal concentration of some
selected antibiotic can sustain to produce the host bacteria stimulation in virulent phase (Silva
and et. al., 2021). The enhancing notion of sub lethal concentration of antibiotic drug for
example name cefotaxime provide the outcome that is observed under increase T4 plaque size
and T4 associated concentration. Technically, synergistic effect is well determining for number
of cases which must be acute or chronic depend on severity (Lin and et. al., 2018). Numerically,
ΣFIC ≤ 0.5 at phage concentration that is associated between 103 and 106 with the unit of
PFU/mL. At the complied level of 1012 PFU of phage’s/mL, the effect was highly additive (≥1
ΣFIC ≤ 2), with this, higher concentration of phage has ability to kill the bacteria without any
antibiotic addition. Non antibiotic alternative treatment are used to show the combat which help
to enhance the infection which is usually causes by multidrug resistant bacteria which steadily
required (Maharjan and et. al., 2022). In some recent years, bacteriophages have remembered to
make the potential replace or complement the role of antibiotic as show the bacterial viruses that
have ability to make the inactivate pathogen. In this, for studying of synergistic effect of phage
antibiotic combination is usually based on some example which contained organism named
Citrobacter amalonaticus and Escherichia coli. These two organism is useful to create
understanding regarding the synergistic effect of phase/anitibiotic to combat the infection. In
reference with Citrobacter amalonaticus isolate of phage antibiotic combination. The Citrobacter
amalonaticus isolate the study which is put together with phage of MRM57 (Narulita and et. al.,
2020). In this, eight different antibiotics with various mechanism of action which is used in
combination with phage to impact the combination treatment on the less inhibitory concentration
or also depend on dilution of organism to understand the resistivity of drug. The research work
usually covers the topic that is well related with synergistic effect of phage/antibiotic as an
effective aspect for combat of infection (Raza, Matuła, Karoń, and Paczesny, 2021).
Antibiotic synergy is defined as response when two or more than two antibiotics are
administered simultaneously with the purpose to treat bacterial infections. Synergistic
mechanism is defined as a process where two or more than two materials produce larger effect in
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comparison to single compound. The antibiotics work collaboratively which enhance the potency
of the drug. The efficacy of the drug also get increased then the single administered drug.
Synergistic effects of drugs are found most effective in order to combat bacterial infections as it
broaden antibacterial spectrum. Administration of combination of drugs ensures that at least one
single active drug will kill or prevent the infecting organism (Zhang and et. al., 2019). For
example, penicillin G is a antibacterial drug and it is combination of gentamicin and
streptomycin. This combination is found effective to deal with 17 and 16 of bacterial strains.
Finding of various peptide antibiotic has leaded in a novel area of research for antibacterial
agents. Ribosomally synthesised natural peptides are discovered as novel therapeutic approach
to treat the bacterial infection due to their antimicrobial potency. Naturally found antimicrobial
peptides are principle elements of the natural host shielding molecules of a very wide range of
plants, species and animals. It has been analysed that naturally found peptides shows broaden
antimicrobial activities (Yang and et. al., 2020). Antibiotic synergy is defined as response when
two or more than two antibiotics are administered simultaneously with the purpose to treat
bacterial infections. Synergistic mechanism is defined as a process where two or more than two
materials produce larger effect in comparison to single compound. The antibiotics work
collaboratively which enhance the potency of the drug. The efficacy of the drug also get
increased then the single administered drug. Synergistic effects of drugs are found most effective
in order to combat bacterial infections as it broaden antibacterial spectrum. Administration of
combination of drugs ensures that at least one single active drug will kill or prevent the infecting
organism. For example, penicillin G is a antibacterial drug and it is combination of gentamicin
and streptomycin shows broad spectrum of antibacterial activity. Studies revealed the
effectiveness of group of natural peptides at the site of action which is cytoplasmic membrane.
This includes mechanism of binding monomers to cytoplasmic membrane in order to form ion
channels pores. There are some multi-drug resistant organism, has been spreading widely and
founded life-threatening for human beings (Gounani and et. al., 2019). Combination therapy of
antibacterial agents helps to treat the bacterial infections effectively. Many research has been
conducted to study the synergy in vivo and in vitro to collect the data about the therapeutic
effects of combination of drugs. Some combinations may result in enhance mutational
intensities. Synergistic drugs have also some negative effects as it increases risk of toxicity and
adverse effect. Research shows that synergistic validity is more effective to prevent antimicrobial
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resistance. Two synergistic drugs are found as effective than single administered drug.
Synergism enhance the efficiency and efficacy of the drug. Multi-drug resistance to treat
bacterial infections is leading across the world (Sharahi and et. al., 2019). Administration of
combination of drugs is considered as an effective novel approach to treat the bacterial diseases.
Synergistic antibacterial activity of group of natural antimicrobial peptides are inactive against
multi-resistant staphylococcal epidermis in vivo and in vitro. Erythromycin, ampicillin and
penicillin are studies as show synergistic effect in combination. The antibacterial activity get
increased with the administration of combination of drugs. Tetracycline drugs shows synergistic
effects with I1WL5W and additive activity with L11W, I4WL5W AND L12W. Since past 10
years, increasing number of multi-resistant bacterial species have enhanced due to
administration of antibiotics across the world. With the help of different mechanism the
development of novel antibiotics has been discovered (Wei, Yu, and Chen, 2019). Synergistic
drugs are recovered after failure of single administered drug. Some bacteria are found drug
resistant and most of the single drug components are not able to kill the infecting bacteria. In
such cases, synergistic drugs are given as increases the potency and efficiency of the drug against
the bacterial infection. Combination of chemicals and active components shows novel approach
to improve and enhance antibacterial activity. Combination therapy or synergistic drugs are
helpful to reduce the dose of the administered drug. It is also helpful to minimize adverse effects
of the drugs. It is a novel approach made to overcome with problems such as adverse effects and
toxicity. Sometimes, combination of drugs are used to overcome the side effects of individual
component present in the drug. Low intensity and high toxicity hinders the development of
antimicrobial agents. Administration of synergistic drugs helps to overcome both of these (Yu
and et. al., 2019).
Aim
To understand the synergistic effect of phage/antibiotics to combat infection.
Objective
 To analyse the notion of synergistic effect of phage/antibiotics to combat infection.
 To evaluate impact of synergistic effect of phage/antibiotics to combat infection.
 To examine the ways and strategies that promote synergistic effect of phage/antibiotics to
combat infection.
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RATIONALE FOR THE RESEARCH
The rationale for the research well show the effectiveness while involving with research work. In
addition, while conducting the research it helps to provide personal as well professional
development. Therefore, the study also generates the knowledge and various skills that is
appropriate as per term of topic. The topic which is chosen for the report is usually focus on the
broad term where investigation is used to done that is about impact of synergistic effect of
antibiotic in combat with infection. The study is broad and provide knowledge that is related
with topic. The personal development means they create sense of knowledge on personal level
which help in future to provide recommendation of antibiotic to relative and friend to combat
with infection. On other hand, professional develop is carried as per the knowledge which is
useful for the future implication when they are related with future placement and courses.
CHAPTER 2: LITERATURE REVIEW (3500)
Notion of synergistic effect of phage/antibiotics to combat infection
With the view point of Prasanth Manohar and colleagues, Antibiotics has been prescribed
to combat infection by resisting the growth or reproduction process of infectious agents within
the body. Specific antibiotics are being prescribed by doctors according to the bacteria or virus
invaded in the body. These drugs eliminate the infectious agents and helps to develop immunity
which assist body to fight against the invaded infectious agents along with natural immunity of
human body, which contains T cells and memory cells. Natural immunity of a human body is too
strong that it can even recognise the infectious agent, if attack further (Pires and et. al., 2022).
The concurrent overuse and over prescription of antibiotics are the main causes of development
of antibiotic resistance in bacteria. Thousands of deaths have been detected every year due to
lack of antibiotic-resistant infections. Shortage of proper and effective treatment options, phage
therapy has re-emerged as a better treatment to treat multidrug resistant infection with
remarkable clinical outcomes. For example; Citrobacter spp. is a gram negative bacteria belong
to Enterobacteriaceae family. These species are commonly found in water, food, soil and the
intestinal tract. They may cause infection in urinary tract, respiratory tract, and bloodstream in
humans. Antibiotics used to treat Citrobacter infections are glycylcycline, aminoglycosides,
beta-lactams, cephalosporins, and polymyxin-B. Beta-lactams is the essential antibiotic class
which target bacterial cell wall. Meropenem, carbenicillin, cefotaxime are highly effective.
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The serious antibiotic crisis has led the researchers to study for the effective of combination of
antibiotics. Even after implementing various approaches to fight against multidrug resistance,
bacteria did not stop to evolve and persist. Phage therapy is a potential option to antimicrobial
chemotherapy, but the use of phages alone without any antibiotics can leads to the development
of phage-resistant bacteria (Chen and et. al., 2022). It has become important to search for an
alternative to stop bacteria from evolving and developing resistance to antibiotics. So antibiotics
and phages have been combined to generate synergistic effect. Phase-antibiotic synergy has
revealed some promising result in eliminating bacteria. Some antibiotics are capable to stimulate
the production of phages through a bacterial host and frame larger plaques along with antibiotics.
And certain antibiotics may have a sublethal concentration which may promote the release of
progeny phages from bacterial cells. Phage-antibiotic synergy (PAS) is so effective that it can
reduce the usage of amount of antibiotic and reduce the emergence of antibiotic resistance in
bacteria. This synergistic combination postulated to be more effective in controlling bacteria.
The research indicates that the combination of antibiotics and phages are more suitable to treat
Gram-positive bacteria including MRSA (methicillin-resistant Staphylococcus aureus) as well as
Enterococcus strains. There is different mechanism of action that can be helpful in explaining the
phenomenon of PAS. 1)cell elongation; 2)size of plaque enhanced by antibiotics, accelerated
plaque amplification and increased burst size; 3)Antibiotic-resistant mutant occurrence decrease
and decrease of phage occur; 4)presence of phage results in increasing antibiotic susceptibility;
5)Antibiotic MIC seems to get reduce due to addition of phages to the antibiotic; 6)Phage
enzyme leads to decompose the bacterial cell wall which facilitate the diffusion of antibiotic and
increase cell penetration (Letrado and et. al., 2018). Comeau and colleagues described that the
process of cell filamentation has been observed during PAS. Quinolones and beta-lactams caused
both PAS and filamentation whereas tetracycline and gentamicin caused neither PAS nor
filamentation. Some researchers said that some antibiotics are capable to promote lysis of cell
given that filamentation induces disturbances in the layer of peptidoglycan and which may
induce phage lysis genes encoded enzymes. This phenomenon can lead to rapid lysis and
enhanced the rate of phage production.
From another mechanism it has been demonstrated that sublethal concentration of
tetracycline, linezolid, and ketolide antibiotics can increase plaque size by three times while
quinolones and beta-lactams antibiotics cannot produce this effect (Tagliaferri, Jansen, and Horz,
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2019). An in vitro study indicated that phage combination that cause infection to S. aureus and
antibiotics (tetracycline, clarithromycin, ciprofloxacin, linezolid, cefotaxime) enhance the
production of progeny phage. Combined therapies of antibiotics and phages were investigated in
vitro to fight against drug-resistant uropathogens. The production of phage MFP through a
uropathogenic E. coli strain can be increased distinctly by low dose of Cefotaxime. It has been
observed that similar effect is effective for beta-lactam, T4-like phages, mitomycin C and
quinolone antibiotics. Lowering antibiotic MIC values give synergistic effects of antibiotics with
E. coli phage cocktail and similar with amikacin when median MIC reduced from 8 to 2 μg/ml
and for fosfomycin, it was reduced from 32 to 8 μg/ml.
PAS have been studied in animal models to evaluate the effect of phages in falling down the
inflammatory reaction, to identify actual synergistic effect of antibiotics and phages. Studies
revealed the data about liposome-encapsulated phages, that they can be effective in diminishing
inflammatory responses. For instance, Kaur and colleagues suggested that levels of TNF-alpha
cytokines, Interleukin-1beta and PCT are surgically implanted in mice by K-wire into intra-
medullary canal, with a specific phage and linezolid antibiotic with lower concentration as in
comparison with K-wire (Luo and et. al., 2022). As a result, intracellular bacterial counts will be
decreased and bring down to a certain level and increasing effect of oxidative killing through
phagocytes. In this situation, the bactericidal activity and phagocytic functions were seeming to
be increased since bacteria are killed and digested by macrophages in vivo. The use and effect of
phages alone as well their combination with antibiotics has also been studied in patients suffer
with suppurative bacterial infection in Poland. To research for the topic of interests, professionals
first administered phages orally or locally and obtained more effective results than the condition
when combined treatment were given along with phages nad antibiotics. Statistically significant
results were observed. From these results, possibility of existence of antagonistic effect between
phages and antibiotics has been considered. An another experiment was conducted by Zilistenau
and colleagues, to check the efficacy of combination of phage-antibiotic and solitary effect of
phage therapy. About 87 patients of chronic UTI has been selected. When phages were
administered orally at 20 ml dose for 5 days and antibiotics were given for 10 days. It has been
observed that almost 92.8% of positive cases shown efficient results while when combination of
antibiotic-phages were given, only 64.4% of patients shown good results. This treatment also
shown the results of existing antagonistic effect between antibiotics and phages.
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Apart from bacterial infection, studies and experiments of combination of antibiotics and
phages has been conducted in case of orthopaedic infections in humans. A trauma-related left
tibial infection patient who has developed the resistance from K. pneumoniae and A. baumanni
has administered the combination of phages K. pneumoniae and A. baumanni through injection
for consecutively 5 days and then after giving break of 1 week, colistin iv and meropenem
administered simultaneously for 6 days observed to give a good clinical outcome with fast
healing of tissues and eradicate positive culture. Such clinical data, it can be evaluated that the
combination of antibiotics and phage M1 caused clinical improvements (Li and et. al., 2021).
Nowadays, phages are used alone by professionals to improve the effectiveness of therapy; by
giving as a supplement for existing antimicrobials or against antibiotic-resistant bacteria. In
humans, cases of recurrent bacterial infections have been treated by applying phages and
antibiotics which shown promising treatment outcomes. Good clinical results have been obtained
in respiratory, orthopaedic, and urinary tract cases from 2018 to 2022 with the application of
combined treatments.
It is important to investigate phage-antibiotic interactions in vitro to intercept antagonism and to
ensure synergistic effect to understand the results of treatments effectively. It should be
considered to understand that phage-antibiotic interaction can depend on dose and type of each
agent as well the time of administration. More research and studies are still going-on to increase
understanding about the combined-action of antibiotics and phages in vivo and in vitro. This may
help in achieving a milestone in near future.
Impact of synergistic effect of phage/antibiotics to combat infection
According to the viewpoint of Deijing, (2019) it is well analysed that the synergistic effect is
required for the various cases where drug is administered in combined form to sum the effect for
better therapeutic effect. Moreover, antibiotic synergy is referring as one of the three process
which is possible when two or more antibiotic are used respectively to the treat the condition or
used to fight against the infection. In the synergistic response, the applied antibiotic is usually
work together to sum the effect which is applicable to reduce the situation which is chronic or
life threatening (Vazquez-Muñoz and et. al., 2019). The applied antibiotic is generally work
together in order to show the effect more potent than if each antibiotic which is were applied. In
this, they are taking into comparison with additive effect, where the potency of an antibiotic
combination which is roughly equal to combined potencies of each antibiotic singly and they
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showing antagonistic effect. Moreover, the potency of combination is less than the combined
potencies of showing each antibiotic (Rezzoagli and et. al., 2020).
According to author of Yan, (2013) the discussion is based on the Gram negative and Gram
positive bacteria which act as the main cause of infection among the mammals. In addition, they
are also showing emergence of their antimicrobial resistance is an enhancing issue in human
medicine. The diabolical fraction or structure which is associated with polycatonic peptide
represent the conserved theme that is based on antimicrobial defence. Therefore, it is essential to
provide new structural class of highly active antimicrobial agents and they are showing the
potential act as resource for development novel anti-infective agents (Namivandi-Zangeneh and
et. al., 2018). Studies is well indicated that they are polycationic peptide that have number of
variable that show antibacterial, antifungal and antiprotozoal activity in vitro chemotherapy. For
instance, compound may be very valuable as adjuvants for antimicrobial chemotherapy. Studies
used to elaborate that to exert their antimicrobial and antiendotoxic activity where peptide must
initiate bind to lipopolysaccharide of outer membrane for gram negative bacteria. The LPGs
usually trigger acute phase response to infection and compound or molecule which is generally
involved in the recognition process that has been extensively studied to show their response for
infection. Context with comparison that is little to known concern towards the bioactive
component which is gram positive bacteria and mechanism through which the target cell is well
activated by such elements (Shahzad and et. al., 2018). Therefore, peptidoglycan is referring as
the major component of the gram positive cell wall and does not contain LPS. Additionally, it is
also analysing to trigger various defensive mechanism as response which is against bacterial
infections. For example, current study confirmed that positive activity of each peptide against
Gram negative or Gram positive bacteria. In addition, the combination effect is well studied that
shows synergy between ranalexin and hydrophobic antibiotic such as polymyxin E, doxycycline
and clarithromycin. Therefore, the magainin II which is well refer as the synergistic with Beta
lactam. In other hand, the mechanism and configuration show the positive interaction which is
appears in the term of complex (Ipe, Kumar, Love, and Hamlet, 2020).
According to the vision of Ekta and Purva, (2022), the synergistic effect is defined with the
help of antibiotic synergy in order to treat infection. In this, treatment of staphylococcal infection
is difficult due to the resistance with the persisted form that pose an added threat of recalcitrant
infection. Therefore, antibiotic combination which is widely studied as an alternative strategy
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with the main aim to treat the condition or to take the combat with the help of synergy that
greater the efficient of drug when they are sum together with same purpose (Wu, Tang, and
Uskokovic, 2018).
Therefore, clinical interest in the synergism dates back to the early of 1950 where healthcare
practitioner noted that patient with the infection for example enterococcal endocarditis are
usually experience high relapse rate when they are treated alone with Penicillin G and
elaborative discussion also configure that lower relapse rate when Penicillin G is combined with
streptomycin to combat with infection. In addition, research community has conducted several of
studies which is about investigation of synergy that is related with drug or medication in order to
treat infection which require additional therapeutic effect to minimise the chronic state of patient
(Chi, and Holo, 2018). The example is well above discussed, in this, the impact of synergy is
well analysed in all the cases where appropriate drug combination such as Penicillin G with
streptomycin show high rate of therapeutic to deal with the infection of enterococal endocarditis.
As per the current time, combination therapy is well recognised as providing the broad spectrum
of antibiotic coverage, effectively combat with poly-microbial infection, reducing selection of
antibiotic resistant strain, minimising the dose toxicity where applicable and in some of cases
they provide synergistic activity (Contreras, Munita, and Arias, 2019).
According to the Zhou, (2022), it is analysed from overall study that antibiotic synergy is
desirable sense for healthcare setting for several. At patient level, the boosted antimicrobial
potency provided by synergy allow the body which is more rapid clear infection that show the
result in shorter courses of antibiotic therapy. Therefore, the shorter courses of therapy which is
showing effect of dose related toxicity. At the higher synergistic effect which is useful for
combating resistant that is based on bacterial strain through enhanced potency and for stalling the
spread of bacterial resistance through the total eradication for the infection. It also shows aspect
of prevention for evolutionary selection of resisted cell and strains (Patrulea, Borchard, and
Jordan, 2020).
Ways and strategies that promote synergistic effect of phage/antibiotics to combat infection
Synergistic effect is referred to as one of the three possible outcomes that occur when more
than two antibiotics are administered simultaneously for the treatment of an infection. The
synergistic effect of antibiotics is also referred to as the antibiotic synergy where the combination
of antibiotics is considered desirable in a clinical sense for various reasons. This condition boosts
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antimicrobial potency at patient level which ultimately results in short courses of antibiotic
therapy. Synergistic effects are also used to combat resistant bacterial strains by increased
potencies and obstructing the spread of bacterial resistance (Grygorcewicz and et. al., 2021). This
is done by total eradication of infection in the body. Antimicrobial resistance is a serious threat to
human race, because of which, the most effective strategy used to fight resistance is through
combination therapy. The use of bacteriophages, in recent years have proven to potentially
replace the role of normal antibiotics. This is because bacterial viruses have the ability to
inactivate or slow the functioning of pathogens. The development of new antimicrobial agents is
crucial to combat infections caused by multidrug-resistant (MDR) bacteria. The combination of
phage-antibiotic synergy is known to promote the process of phage replication and reduce the
growth and development of bacterial resistance. The antimicrobial efficacy and the therapeutic
performance of phage-antibiotic synergy is known to positively impact the bacterial inhibitory
effects to extreme levels. The rise of antibiotic resistance is the reason why phage therapy has
been increasingly studied and researched upon (Gonzalez‐Moreno and et. al., 2021).
The antimicrobial action produced from the combination of bacteriophages and antibiotics
should be promoted and the information about this study should be distributed so that all patients
can shift from administration of conventional antibiotic medications. The main focus for the
promotion of antibacterial activity should be evidence based (Chang, Nang, Chan and Li, 2022).
The use of antimicrobial phage therapy is essential to be evidence based because of its
application on human body. The application of proper research methods in wide range of fields
such as biology, chemistry, toxicology and pharmacology is crucial during the development of
new medicines. Without the application of clinical research, there is no evidence to prove the
efficacy of new treatments over the old ones. Therefore, research overview is one way through
which healthcare workers navigate towards finding the most effective therapy for care for
patients. Application of appropriate research methods towards identification of efficacy of
bacteriophage-antibiotic therapy information which is reliable and authentic. Along with
implementation of research methods, for the promotion of synergistic effect of combination
therapy, it is crucial to build credibility of information. The emphasis will be laid more on the
principles of the therapy rather than invalid and inappropriate information. Along with
knowledge build-up, the appropriate research about this therapy will ensure the understanding of
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