**5.9 Tuberculosis**

Tuberculosis (TB) is a lung infection caused by the endogenous bacterium *Mycobacterium tuberculosis*. First-line TB drugs drugs like rifampicin and isoniazid are resistant to certain types of multidrug-resistant tuberculosis (MDR-TB). In 2018, 484,000 new TB patients failed to respond to rifampicin, according to the World Health Organization (WHO). Seventy-eight percent of these patients have tuberculosis with multidrug resistance (MDR-TB) [118]. Mycobacteria exist in over 170 distinct species, each with its own pathogen development in humans [119]. *Mycobacterium ulcerans* and *M. leprae*, in addition to tuberculosis, M*ycobacterium* ulceration and M*ycobacterium* leprosy, respectively, cause Barley ulcers and leprosy [120]. Alternative therapies for MDR-TB are important for disease control, particularly as newer approaches to mycobacteriophage therapy emerge. To date, 11,282 mycobacteriophages have been discovered [121]. *M. smegmatis*, a non-pathogenic vector, can transport phages to the same intracellular compartments as *M. tuberculosis* [122]. M. mycobacteriophage D29's antimicrobial value was doubled after it was given twice in a 24-hour period to treat tuberculosis H37RV [123]. Aerosolized bacteriophage D29 treatment reduced TB cases in the lungs and vaccinated mice against tuberculosis [124]. Tuberculosis-prone health workers can benefit from aerosolized mycobacteriophages. D29 was used to treat burly ulcers caused by Mycobacterium ulcers in the Marine Footpad model [125]. As the disease progresses, infected patients experience necrosis of the skin, subcutaneous tissue, and bone, necessitating surgical skin rupture. Mycobacterial and pathological counts were decreased after D29 was injected subcutaneously. In footballs and lymph nodes, it causes the development of water-borne cytokines. This approach was used to deliver the lytic mycobacteriophage TM4 to *M. tuberculosis*-infected RAW264.7 macrophages, which decreased bacterial counts. On the other hand, the phage was found to be inactive on its own. *M. smegmatis*-TM4 complex substantially reduced bacterial counts in *M. avium*-infected mice's spleens, while TM4 or *M. smegmatis* alone had no effect [126]. Phage cocktails may be used to overcome phage resistance tuberculosis.

### *5.9.1 Endolysin therapy*

Endolysin therapy is a major part of phage therapy. Endolysins are considered protein-based antibiotics or antimicrobials. The purified endolysin is a powerful antibacterial agent for curing bacterial infections in human beings and animals. The efficacy of endolysin enzyme, host bacteria, bacterial disease and the therapuetic use in experimental animal models is listed in **Table 6**. Endolysin is an enzyme used by the bacteriophages to degrade the bacterial host's peptidoglycan from the inside,





**Table 6.**

 *Endolysin therapy: therapeutic effects of Enzymes on bacterial diseases.*

resulting in the release of cell lysis and offspring virions [23, 115]. *In vitro* and in mice models, the recombinant phage-derived lysins exhibit highly efficient bactericidal activity against multidrug-resistant *E. faecalis*. Endolysin LysEF-P10, EF24C, Lys168, Lys170 PlyV12 LysEF-P10, IME-EF1, and lysine CF-301 zap methicillinresistant *Staphylococcus aureus* (MRSA). Antibiotic-resistant *S. pneumoniae*/Acute otitis media (AOM) infected mice exhibited a quick recovery from infection after 24 hours when they were treated with CPL-11 (therapeutic pneumococcal lysin streptococcal bacteriophage) at a dose of 2,000 μg [117]. The mixture of lysostaphin and the chimeric phage lysin λSA2E-LysK-SH3b synergistically kill *S. aureus in vitro* and in mouse models of bovine mastitis [140, 141]. Some lysins, such as CF-301, N-Refasin, P128, and Art-55, are at various stages of pre-clinical or clinical development and are antibacterial for the cure of multiple antibiotic drug-resistant (MDR) infections of Gram-positive, and Gram-negative pathogens. Synchronization of pneumococcal phage lysine with CPL-1 and autolysin LytA eliminates *Streptococcus pneumonia, S. pseudopunemonia,* and *S. aureus* [80, 140]. Endolysin shows synergistic action with phage lysin LySMP or antibiotics that are very specific to cell wall components and is considered an alternative to drug antimicrobial therapy because lysine kills target bacteria rather than other microorganisms. There has been a significant increase in potential applications of phage lysin which is specifically promising, kind of topically applied therapeutics, and lysin also becomes a practicable alternative to antibiotics in long-term systemic therapy [117, 142, 143]. Endolysins have been used effectively in medical applications. They exhibit specific antimicrobial activities in controlling and treatment of pathogenic bacteria such as *Streptococcus* and *Staphylococcus*. Beneficial synergistic interactions increase the efficacy of treatments and reduce the risk of resistant strain development. There was no inactivation nor adverse side effects detected *in vivo*. The creation of chimeric proteins by rearrangement of functional domains of lysins of multiple species established molecular engineering of lysins which can increase lytic activity, widen specificity, advance binding affinity, enhance solubility and reduce the chance of resistance formation, thereby optimizing lysins for specific applications. Moreover, endolysin-based antimicrobials viz. (Outer membrane permeabilizers (OMPs) and protein transduction domains (PTDs) are used to control Gram-negative and intracellular pathogens. Molecular engineering of lysins is predicted to gain momentum in the coming years and the dogma of endolysins to be effective only against Grampositive bacteria when applied externally to decrease [144].

### *5.9.2 Side effects*

Most of the drug resistance in bacteria studies had been carried out on the use of *in vitro* and *in vivo* experiments in animal species with a particular look upon human studies. Predicaments of the misuse of bacteriophages as medicament specialists are in many situations recognized into four classifications: (1) phage selection, (2) bacteriophage have host-range restrictions, (3) the uniqueness of phages as recommended medications, and (4) uncommonness with phage. Studies on T4 phage had recognized no massive fitness effects and pronounced negative results such as inconvenience, itching, wetness, and unattractive scent, unfavourable events, sore throat, belly pain, nausea, extended peristalsis [65, 66]. *Staphylococcus* bacteriophages proved drug intolerance and hypersensitive manifestations at the site of injury on days three to five of bacteriophage therapy, and hepatalgia was detected after several hours [145, 146]. Adverse activities occurred in six (21%) of 28 victims in the Pyophage team in distinction with 13 (41%) of 32 victims in the placebo group Urinary tract infection intravesical Pyobacteriophage (Pyophage;

20 mL) [112]. A cocktail of 12 lytic anti-*P aeruginosa* bacteriophages in the PP1131 group, twenty-three (23%) of 13 analysable individuals had adverse reactions versus seven (54%) of 13 in standard-care-group [83]. It is gratifying to understand renewed interest in bacteriophages which is nature's different tailored solution to the problem of antibiotic-resistant bacteria. Beyond the urgent problem of untreatable infections, detailed research of bacteriophages have the possibility of finding, exhilarating new biology, molecular mechanisms of RNA-guided DNA targeting and cleavage by the Cas9 enzyme Cas9 in genome engineering effective use of CRISPR-mediated understanding of CRISPR–Cas9 mechanisms genome engineering in clinical applications CRISPR biology exemplified via the transformative discovery of CRISPR–Cas DNA editing structures and phage-encoded anti-CRISPR defences [51]. We're on the verge of entering an exciting new era in phage technology and applications, thanks to advanced molecular methods, devices, and applications in medicines.
