Mycoplasmas are potential pathogenic organisms of humans and many animal species. Pathogenic mycoplasmas have a natural tendency to colonize certain sites in vivo called “tissue tropism”, such as respiratory, ocular and genital mucosa, and mammary glands (Razin et al., 1998)
Human Infection
Mainly, six Mycoplasma species (M. pneumoniae, M. genitalium, Ureaplasma (U) urealyticum, U. parvum, M. hominis, M. penetrans) have been demonstrated to cause human illness (Waites et al., 2005), such as acute respiratory disease (Waites et al., 2017), joint infections (Ali et al., 2021a; Mărginean et al., 2021), genital and urinary tract infections (Peter et al., 2018), and neurological disturbance (D’Alonzo et al., 2018; He et al., 2021a). On the other hand, some species mainly infecting animals like M. suis, M. ovis, and M. haemofelis have been detected in humans and regarded as zoonotic pathogens (Maggi et al., 2013b). These mycoplasmas can establish persistent infections (Yavlovich et al., 2004), alter host cell physiology, modify apoptotic pathways (Chernov et al., 2015), induce the production of inflammatory substances (Benedetti et al., 2020), and result in cellular DNA damage and cancers (Zella et al., 2018). In addition, serious consequences like chronic obstructive pulmonary disease (COPD) (Feng et al., 2021) and infertility may occur (Kusanovic et al., 2020).
Transmission of human Mycoplasma infection occurs through human-to-human contact. It mainly infects mucosal surfaces of the respiratory and urogenital tracts. Droplets containing the organism spread the infection from host to host (Waites et al., 2017). Ureaplasma species, M. genitalium, and M. hominis are genitourinary mucosal organisms and the infection can spread through direct sexual contact, Ureaplasma species mainly cause urethral and gynecological infections (Cassell et al., 1993; Lanao et al., 2022).
The clinical picture of M. pneumoniae has diversely presented from self-limiting to life-threatening disease (Saraya et al., 2014). For instance, it causes community-acquired pneumonia (CAP) in people of any age, especially in children and young adults (Li et al., 2019; Tsai et al., 2021). M. pneumoniae symptoms are variable including fever, cough, sore throat, and occasionally, acute exacerbation of asthma. In addition, severe pulmonary sickness including bronchiolitis, pleural effusion, lung abscess, and pulmonary embolism as a consequence of M. pneumoniae infection has also been reported (Meyer Sauteur et al., 2016). The culture procedure is a gold standard technique for M. pneumoniae diagnosis. On the other hand, it is recommended to use polymerase chain reaction (PCR) for diagnosing acute cases because culture methods require several days for obtaining results. Moreover, PCR is a rapid, sensitive, specific, and commercially available method, and so it is more suitable for mycoplasmas diagnosis in the clinic (Waites et al., 2017).
Mainly, macrolides, fluoroquinolones, and tetracycline are used for the treatment of M. pneumoniae infection; macrolides are the most potent antimicrobial agents for the treatment of mycoplasmosis through inhibition of the bacterial protein synthesis (Waites et al., 2017). Recently, because of the broad use of macrolides, macrolide-resistant M. pneumoniae (MRMP) has become increasingly prevalent worldwide (Yang, 2019). The macrolides resistance of M. pneumoniae has been emerging worldwide. In Taiwan (2010 to 2017), its rate was 15–30% (Yang et al., 2019a). In America and Europe (2008 to 2013), it was <30%. While in other countries and regions (China mainland, Japan, and Korea) it was about 60–90% (Waites et al., 2017). Age is also regarded as a major determinant for MRMP as the detection rate was higher for children aged ≤15 years than adults. In adults, the detection rate was higher in adolescents (16–19 years) than in older age (≥20 years) (Yamazaki and Kenri, 2016). On the other hand, fluoroquinolones and tetracyclines have more severe side effects than macrolides as tetracycline cause enamel hypoplasia and discoloration of the teeth in young children. Despite the detected hazardous effects of fluoroquinolones on joints and muscles of children, they have been successfully used for the treatment of some complicated cases of MRMP strains in young children (Jackson et al., 2016; Ahn et al., 2021).
Owing to the marked increase of M. pneumoniae antimicrobial resistance in recent years as well as the previously mentioned side effects of some antimicrobial agents, the development of protective vaccines against this pathogen is a critical requirement (Jiang et al., 2021). Recently, the designing of the next-generation vaccine approach was performed to establish an effective multi-epitope vaccine (MEV) for human protection against M. pneumoniae (Mahmood et al., 2021). To date, Mara et al. (2020) have achieved a breakthrough in explaining how the vaccine-enhanced disease (VED) occurs as a result of M. pneumoniae vaccination with lipid-associated membrane proteins (LAMPs). Intriguingly, they demonstrated that M. pneumoniae lipoproteins lipid moieties are responsible for VED occurrence. In addition, the removal of lipid molecules from LAMPs before vaccination prevents VED and reduces bacterial loads in the case of M. pneumoniae infection. Lipoproteins are the main immunogenic and antigenic constituents of the LAMPs fraction, and therefore their lipid moieties significantly reduced LAMP-stimulated TNF-α production which leads to the VED (Mara et al., 2020). These results may be widely applicable for other mycoplasmas in which vaccine-induced disease exacerbation has been described such as M. bovis (Bryson et al., 1999) and Mmm (Nicholas et al., 2004).
Nowadays, sexually transmitted antigens are of major concern. Nogueira and co-workers have conducted a recent computational study using in silico methods as “subtractive genomics and reverse vaccinology” on five strains of M. genitalium, a serious sexually transmissible pathogen. The state-of-the-art sequencing technologies with the availability of the required genomic data paved the way for conducting this work that aimed at predicting the potential vaccine targets and drug candidates. A total of 14 novel vaccine candidates and 2 novel drug targets have been obtained which need further experimental validation to ensure their efficacy for the prevention and control of M. genitalium infection (Nogueira et al., 2021). More interestingly, M. genitalium is resistant to most antibiotics and difficult to be treated and controlled. Also, it causes endometritis, premature birth, and sterility in women and urethritis in men (McGowin and Totten, 2017). Hence, Ali et al. have conducted proteome-wide vaccine targets prioritization for designing an antigenic vaccine candidate against M. genitalium infection. MEV has been constructed successfully with further determining of the different physicochemical properties of the vaccine, but this study still needs further experimental validation for the constructed MEV (Ali et al., 2021b).
More recently, M. hominis infection was reported to cause bacteremia, pneumonia, and meningitis, but its significance to cause neonatal meningitis remains elusive. Using CSF patient samples, translucent colonies were observed on chocolate agar media, and the microorganism was recognized as M. hominis with MALDI-TOF MS. The 16S rRNA gene sequencing was also carried out which showed 99% nucleotide identity to M. hominis (Kersin et al., 2020). M. hominis is characterized by a very slow growth rate that requires specific growth media and it’s resistant to many antibiotics such as β-lactams, glycopeptides, sulfonamides, and macrolides (Kersin et al., 2020; Ferreira et al., 2022).
Mycoplasma Infection and Respiratory Diseases
Traditionally, the clinical picture of Mycoplasma infections was more intimately suggestive of damage due to host immune and inflammatory responses rather than direct toxic effects induced by Mycoplasma cell components (Razin and Jacpbs, 1992). However, He et al. (2018) have shown that many direct effects, as well as indirect immune mechanisms, have been incorporated in M. pneumoniae pathogenesis. The direct effect mechanisms include adhesion damage of M. pneumoniae to targeted epithelium then membrane fusion damage via alteration in its exposed receptors (Bao et al., 2015). In addition, nutrition depletion is caused by its limited metabolic capacity (Yus et al., 2009). Invasive and toxic damages are following mycoplasmas invasion of different host cells and microbial production of H2O2 and superoxide. Besides, the produced endogenous toxic oxygen leads to an increase in the intracellular oxygen pressure in the host cells, subsequently; oxidative stress and cell death will occur (He et al., 2018) as shown in Figure 1. On the other hand, the indirect immune damage mechanisms include humoral and cell-mediated damages and inflammatory damage via an intracellular receptor protein complex (inflammasome) (Shimizu, 2016).
More recently, gene expression analysis and whole transcriptome sequencing have been performed for M. pneumoniae infected Hela cells. The results illustrated that protein-coding genes of M. pneumoniae are correlated with immune response rather than cellular processes, probably suggesting the intrinsic ability of M. pneumoniae to modulate host immune pathways (Ramos et al., 2021).
COPD is one of the foremost predisposing causes of death in the USA, killing > 130,000 individuals per year. Globally, > 3 million deaths annually of COPD (Marciniuk and Schraufnagel, 2017), meanwhile, the middle- and low-income countries are more severely affected. Moreover, the lung microbiota of COPD patients contains more M. pneumoniae (Marciniuk and Schraufnagel, 2017). Eventually, M. pneumoniae continues to significantly aggravate the onset and recurrence of asthma (Meyer Sauteur et al., 2016).
Mycoplasma Infection and Urogenital Diseases
In females, M. genitalium, a sexually transmitted pathogen, has been associated with cervicitis, pelvic inflammatory disease (PID), spontaneous abortion, preterm delivery, and infertility. In parallel, it was detected among 10% to 30% as well as 4% to 22% of women with clinical cervicitis and PID, respectively (Gaydos et al., 2009). The high susceptibility of emerging antibiotic resistance is becoming increasingly important (Peter et al., 2018). On the other hand, M. genitalium causes symptomatic and asymptomatic urethritis among men and is the etiology of approximately 15%–20% and 40% of Nongonococcal urethritis (NGU) and persistent or recurrent urethritis, respectively (Bachmann et al., 2020).
Additionally, most urinary tract infecting bacteria can be demonstrated on standard culture, but it is exceptional for Mycoplasma and Ureaplasma. In which, bacterial count in urine does not necessarily relate to the number of bacteria in the bladder wall. A significant number of these intracellular organisms may occur in the bladder wall and be absent in urine. Thereafter, unresponsiveness to antibiotics, persistent lower urinary tract infection, and pyelonephritis were previously reported (Combaz-Söhnchen and Kuhn, 2017). M. hominis and Ureaplasma infection is notably associated with women’s infertility (Latino et al., 2018; Kusanovic et al., 2020). A recent study has identified these infertility-causing pathogens using the PCR technique. A number of 2360 tissue samples have been collected by urethral and cervical canal scrapings of adult women suffering from PID. The results showed that Ureaplasma spp. and M. hominis have been identified in 543 and 179 women, respectively. In addition, 112 women had mixed infections (Piscopo et al., 2020).
Furthermore, M. hominis, U. urealyticum, and U. parvum are examples of pathogens that can invade pregnant mothers and are closely associated with neonatal pneumonia (Waites et al., 2005). In pregnant mothers, U. urealyticum is found in the lower urogenital tract flora, occasionally, it ascends and causes bacterial vaginosis, chorioamnionitis, and premature birth (Stol et al., 2021). In the fetus, it causes neonatal sepsis and meningitis (Ferreira et al., 2021). Both Ureaplasma spp. and M. hominis can produce spontaneous abortion with higher rates in the case of M. hominis (Latino et al., 2018; Kusanovic et al., 2020). Genital mycoplasmas and ureaplasmas can colonize the urogenital tract which leads to invasive infection and spread to the placenta (Huber et al., 2018). Also, congenital M. pneumoniae pneumonia may take place via invasion and hematogenous transplacental infection (Hooven and Polin, 2017).
Mycoplasma Infection and Joints, Blood, Neurological, and Bone Disorders
M. pneumoniae has been frequently involved in severe CNS diseases, such as encephalitis (Al-Zaidy et al., 2015). Besides, it is associated with acute transverse myelitis (ATM) in the form of acute bilateral lower extremity paralysis, paresthesia, and bowel and bladder dysfunction. This syndrome was observed in 15-year-old patients with a slow curing rate that paid attention to the importance of early identification of mycoplasmas infection as a causative agent of ATM and more severe neurological complications (He et al., 2021a). Several extrapulmonary lesions such as cardiovascular, digestive, musculoskeletal, and dermatological lesions during M. pneumoniae infection have been summarized in a mini-review reported by (Narita, 2016).
Clinically, arthritis associated with M. pneumoniae infection has been diagnosed in children (Azumagawa et al., 2008). In addition, septic arthritis caused by M. hominis (Ali et al., 2021a) as well as U. parvum (MacKenzie et al., 2010) has been reported in immunosuppressed patients. M. hominis has been identified as a novel periprosthetic joint infection (a rare postoperative complication) using a new tool called “metagenomic sequencing” (Wang et al., 2021a). M. hominis also might cause brain abscesses (Whitson et al., 2014). What’s more, Ureaplasma species have been reported as pathogenic agents causing CNS inflammation in premature babies and abscessation in the adults’ brains (Glaser and Speer, 2015).
Infrequently, M. orale, an organism that is generally considered non-pathogenic in humans, has been isolated from patients with immunodeficiency and, as a result, these patients suffered from multiple abscesses and destructive bone disease. For direct detection of the pathogen, surgical specimens were used to do 16S rRNA sequence analysis (Paessler et al. 2002). Recently Ketchersid et al., have described a case report of recurrent multifocal M. orale infection in an immunocompromised patient (Ketchersid et al. 2020). Eventually, hemotropic mycoplasmas (HM) are epierythrocytic pathogens that attach to red blood cells of various mammals, including humans causing severe hemolytic anemia (Maggi et al., 2013b; Ikeda et al. 2017).
<italic>Mycoplasma</italic> Infection and Cancers
In recent years, many scientists carried out in vitro studies using oral tissues (Patil et al., 2015), hepatocytes (Choi et al., 2014) cervical cells (Atallah et al., 2020), and human prostate cells (Abdul-Wahab et al., 2021). These studies concluded that mycoplasmas infection stimulates tumorigenesis by inducing cellular transformation. Yacoub et al. (2021) have demonstrated the possible relationship between mycoplasmas infection and the development of cancers. In parallel, they reported the induction of malignant transformation by mycoplasmas infection in PMNCs (Zhang et al., 2004) and in many other human cell lines such as the uterus SK-UT-1B cells (Polianskaia et al., 1998), A549 lung cells and bone tissues (Jiang et al., 2008), prostate BPH-1 cells (Namiki et al., 2009), and neuronal cell lines (Ji et al., 2019). Another study used the PCR technique to determine that M. genitalium levels in patients with prostate cancer were significantly higher than those of patients with benign prostatic hyperplasia (Namiki et al., 2009; Barykova et al., 2011). As such, the available in vitro experimental data indicate that Mycoplasma infection induces chromosomal alteration, chromosomal instability, and/or cellular transformation via genetic mutations and translocations (Paton et al., 1965; Tsai et al., 1995; Feng et al., 1999; Zhang et al., 2000). M. penetrans, M. fermentans, and M. hyorhinis have been observed to stimulate chromosomal abnormalities, which in turn alter gene expression and cause malignant cell transformation (Benedetti et al., 2020). In addition, M. hyorhinis induces hepatocellular carcinoma (HCC) cell migration via the interaction of p37 protein with an epithelial cell adhesion molecule (EpCAM). P37 protein plays a key role in facilitating metastases and invasiveness of various cancer cells (Kim et al., 2019). Benedetti et al. have described Mycoplasma chaperone DnaK protein as responsible for cellular transformation. Besides they substantiated that this chaperone protein binds to Poly-(ADP-ribose) Polymerase (PARP)-1, a protein that is involved in the repair of any possible DNA damage, and reduces its defensive action. It also binds to USP10 which acts as an essential regulator for p53 protein and minimizes the p53 anti-cancer functions (Benedetti et al., 2020). Furthermore, using the in vivo mouse model, it was stated that specific-pathogen-free (SPF) conditions reduced the possibility of tumors formation. Therefore, the diverse microbiome compositions with predominant intracellular mycoplasmas affect the association between the diverse species of Mycoplasma and human cancers (Huang et al., 2001; Pehlivan et al., 2004; Pehlivan et al., 2005). In other words, mycoplasmas have been found in many tumor types. So, it is important to identify and characterize the mycoplasmas associated with the tumors in order to determine their role in carcinogenesis (Goodman and Gardner, 2018).
The first report of empyema caused by a commensal human Mycoplasma infection was described in a case of right pleural space infection with M. salivarium that was accompanied by laryngeal cancer (Baracaldo et al., 2012). M. salivarium is commonly responsible for nonpathogenic human infections, but it causes pathogenic infections only in the case of immunosuppressed persons through invasion of the human oropharynx (Totten et al., 2021).
Animals Infection
Many species of domestic and wild animals suffer from mycoplasmosis. Of which, contagious bovine pleuropneumonia (CBPP) and contagious caprine pleuropneumonia (CCPP) are the two most serious diseases, especially in low and middle-income countries shown by pleuropneumonia accompanied by extremely painful symptoms, reduced productivity, and death. CBPP has higher morbidity compared with CCPP; however, CCPP has higher mortality rates (Bolajoko et al., 2020). Both diseases require accurate diagnostic techniques and improved vaccines which should be accessible in the affected countries (Jores et al., 2020).
Bovine Infection
CBPP is mainly a disease of cattle and water buffalo, it is caused by Mycoplasma mycoides subsp. mycoides (Mmm) and notifiable disease of cattle listed by the World Organization for Animal Health (OIE) (Grieco et al., 2001; OIE, 2021). For the time being, OIE has announced that Europe, the USA, Australia, and South Africa are free from CBPP. For Asian countries, China and India were declared to be officially free, but the disease status is currently unknown in the remaining parts of Asia (OIE, 2019). However, it is endemic in sub-Saharan Africa causing huge annual economic losses (almost 2 billion US$), high mortality (10-70%), severe fibrous bronchopneumonia in the acute cases, and pulmonary sequestra in the chronic stage (Anonymous, 2018).
Mmm infection can be summarized in several consecutive stages, firstly, inhalation of infected aerosol droplets; after that, colonization of bronchioles and alveoli, thereby; Mmm invades the blood and lymphatic vessels and causes vasculitis. Finally, Mmm passes through blood and persists in a variety of other tissues including the lung, in which, the antigen is mainly detected in lung phagocytic cells, on the alveolar and bronchiolar epithelial cells, within the wall of blood and lymphatic vessels, and inside necrotic areas (sequestra formation) as shown in Figure 3. Infected animals actively excrete the pathogen through aerosolized droplets as a potential source of infection for the closely in-contact animals (Di Teodoro et al., 2020). The attenuated CBPP vaccine can provide a moderate level of protection estimated by a reduction in lung lesions in vaccinated and challenged cattle. Annual revaccination with the live vaccine is necessary to maintain protective immunity. Additionally, this vaccine is relatively inexpensive and easy to be produced on a large scale (Jores et al., 2020). On the other hand, this vaccine has a short period of immunity with many adverse reactions because it is a live-attenuated type; therefore, its reversion to virulent form sometimes occurs. In addition, it is temperature-sensitive (Dudek et al., 2021). Often, severe inflammation at the injection site followed by skin sloughing has been reported, and so far it can lead to animal death (Jores et al., 2020).
M. californicum, M. leachii, and M. dispar are other mycoplasmas that can cause significant diseases in cattle, but the most important worldwide pathogen infecting cattle is M. bovis. It can quickly spread to all age groups. Newborn calves can get the infection from older animals that suffer from severe mastitis, arthritis, and pneumonia (Hazelton et al., 2020) that maintain the infection cycle in the herd. Following a recent survey conducted in the United Kingdom from 2006 to 2017, calves at the age of < 3 months (post-weaning) have the highest prevalence of M. bovis pneumonia Mycoplasma bovis Investigations in Cattle (2018). Mainly, M. bovis infects the upper respiratory tract of young calves during the first few weeks of life through feeding of infected milk and/or direct contact with other infected calves’ nasal secretions (Maunsell et al., 2009). Hence, to stop the infection chain, we must stop the infection spread to the new calves born after the M. bovis detection on the farm. Methods for controlling M. bovis are culling or isolating M. bovis mastitic cows, pasteurization of infected milk, raising the calves separately from older animals, and better milking hygiene and teat dipping. But unfortunately, until now the protective vaccine against this serious pathogen is commercially unavailable (Haapala et al., 2021). Moreover, it is one of the four main bacterial pathogens associated with bovine respiratory disease (BRD) with significant economic losses as a result of higher morbidity and mortality rates, reduced growth performance, and raised costs of prevention and treatment (Kudirkiene et al., 2021).
Caprine Infection
CCPP is a fatal contagious illness of goats caused by Mycoplasma capricolum subspecies capripneumoniae (Mcc) and a notifiable disease listed by OIE. It has been reported to affect wild and domestic caprines. A recent report has estimated that CCPP has different case fatality rates of 30% in goat herd (n=200) and 8% in sheep flocks (n=400) (Abd-Elrahman et al., 2020) though previous workers have found sheep to be far more resistant (Nicholas et al., 2008). In addition, It is more widely endemic in East Africa, particularly in Kenya, Tanzania, and Ethiopia (Falquet et al., 2014). The first step to establishing a successful vaccine of CCPP is to design a challenge model that can be used to perform essential immunological studies. As this microbe proved to be host and tissue-specific, a novel challenge model has been established following the recent Kenyan outbreak strain ILRI181 in 2012 (Falquet et al., 2014) rather than the old Kenyan strain F38 (MacOwan and Minette, 1976). The base of this model is using two consequent inoculations of aerosols of Mcc culture into the nasal cavity of goats than a trans-tracheal inoculation of animals. This model has a morbidity of 100% and a mortality of 50–60% which simulate the natural infection pattern (Liljander et al., 2019). The current CCPP vaccine is a bacterin with saponin adjuvant, scheduled to start vaccination at 4 months of age with revaccination every 6 months. It is expensive due to the fastidious growth requirements of the pathogen and the relatively high total protein required for one dose of the vaccine (Jores et al., 2020).
M. agalactiae is a causative agent of an OIE notefiable disease called contagious agalactia (CA) that causes mastitis in dairy goats with formidable financial losses due to arthritis and drop or complete cessation of milk secretion, cachexia, and cornea opacity that can give rise to complete blindness (Santos et al., 2015). In Brazil, the estimated prevalence of CA in goats in different Brazilian provinces such as Rio Grande do Norte, the main goat raising state, was 83.28%, São Paulo was 27.7%, (Azevedo et al., 2015) and Sergipe was 10.3% (Santos et al., 2015; Damasceno et al., 2020). More importantly, many Mediterranean countries are showing substantial losses in the goat dairy industry in France (Poumarat et al., 2016), Spain (Paterna et al., 2013), and Italy (Cillara et al., 2015). CA causes considerable economic losses in Ukraine, according to a recent serological investigation in the Artsyzk area, 168 ewes (32.4 percent) of 519 investigated animals were infected with contagious agalactia. Of which, 109 (64.9%) were in their first year of life, 52 (31.0%) in their second year, and 6 (3.6%) in the 5-6-year-old age group (Bohach et al., 2021).
Hemoplasmas are known as pleomorphic tiny bacteria; they were named because they tend to attach to the erythrocytes’ surface and may cause hemolytic anemia in a wide range of mammals as well. Two well-known hemoplasmas, M. ovis and Candidatus M. haemovis, have been proven to infect small ruminants, their severity increases in young aged and pregnant animals (Hornok et al., 2012). M. ovis is a causative agent of chronic infection in caprines. Few reports are available regarding the prevalence of M. ovis infection in goats with variable figures ranging from absence in Australia and Tunisia (Rjeibi et al., 2015) to 20% in Hungary and 94% in Malaysia (Jesse et al., 2015).
Ovine Infection
Since M. ovipneumoniae was isolated for the first time; it is widely known as “sheep atypical pneumonia” specifically infecting sheep and goats (Besser et al., 2013). Latterly, it causes a potential threat to fattening lamb flocks and the lamb industry due to lower lamb growth and decreased ewe productivity rates, it also has been reported in many worldwide epidemics (Bai et al., 2020) (Jaÿ et al., 2020).
Urie et al. had conducted a wide-scale study and estimated the overall prevalence of M. ovis infection across the USA, it was 24.3% in domestic sheep (Urie et al., 2019). In another study, M. ovis prevalence was high up to 45.8% in 504 sheep samples in China (Wang et al., 2017). Importantly, Maggi and coworkers have reported that M. ovis-like species was the most predominant hemotropic organism found in human patients; thus, M. ovis could have a zoonotic nature (Maggi et al., 2013b).
Swine Infection
M. hyopneumoniae (M. hyo) and M. hyorhinis (Stemke et al., 1992) have been recognized as the main Mycoplasma species that are responsible for various porcine respiratory disorders. Merodio et al. have applied an experimental swine infection model of M. hyorhinis, the results indicated that multiple inoculations may simulate subclinical natural infection as in the field. Besides, animals would have to be infected several times for showing a visible immune response (Merodio et al., 2021). M. hyorhinis and M. hyosynoviae, are commensal microbes of the upper respiratory tract and tonsils of swine, they cause arthritis and polyserositis in young pigs between (6-10) weeks of age. While pigs older than 3 months of age are usually suffering from mild arthritis (Neto, 2012). More frequently, M. hyosynoviae is known to cause arthritis in adult pigs, but its lesions are restricted to the joints and synovial membranes (Gomes Neto et al., 2015).
M. hyo plays a significant role in the development of the porcine respiratory disease complex (PRDC) infection via reduced animal growth performance, reduced feed efficiency, and decreased average daily gain. Mostly, an increase in mortality rate takes place with the help of complicated infections (Pasteurella multocida, Haemophilus parasuis, Streptococcus spp., and Actinomyces pyogenes) which leads to increased total fatality rates (Olaniyi et al., 2020). Multilocus variable-number tandem repeat analysis (MLVA) and multilocus sequence typing (MLST) are strain typing genetic tools that can be used for M. hyo diagnosis (You et al., 2020). A recent study concluded that the most significant histological changes recorded were thickening of alveolar septa caused by neutrophilic cellular infiltration with intraluminal cellular exudate. The majority of pulmonary lesions were chronic (75.81%) (Mucha et al., 2020). Gilts are considered the main source of pathogen inlets because they are mostly exposed to the pathogen during the lactation period (Patterson and Foxcroft, 2019). Vaccination is frequently administrated all over the world with various commercially available M. hyo vaccines for not only healthy animals but also infected herds (Maes et al., 2020). For controlling M. hyo infection, Sponheim et al., have recommended the deep tracheal catheter as a more sensitive sampling tool, used for M. hyo diagnosis, than laryngeal swabs (Sponheim et al., 2020).
Another threat to the pig industry is infectious anemia caused by three hemoplasma species, M. haemosuis, M. suis (Eperythrozoon suis), and Eperythrozoon parvum. M. suis is the main causative agent of swine hemoplasmosis, which in turn adheres to the RBCs surface and triggers their engulfing by the spleen (Petri et al., 2020), as well as causes reproductive failure mainly stillbirths as reported in Southern Brazil (Bordin et al., 2021).
More importantly, M. suis has been proved as the first member of the HM group able to invade the erythrocytes of its host. Using electron microscopy, Groebel et al., have discovered a novel M. suis invasive strain that causes severe swine anemia with a fatal illness. Such invasion enables it to escape the host’s immune response and antibiotic therapy, and the intracellular lifestyle has clarified the chronic nature of HM infections (Groebel et al., 2009). Moreover, the genus Eperythrozoon was previously transferred to the genus Mycoplasma. Now it’s classified under a new order called Mycoplasmoidales (Gupta et al., 2018).
M. haemosuis was associated with fever, anemia, and skin lesions in domestic pigs (Stadler et al., 2020). Genus Eperythrozoon has two new blood parasites species (Eperythrozoon suis and Eperythrozoon parvum), and it was associated with a severe swine disease called “anaplasmosis-like disease” (Splitter, 1950). Globally, the recently detected swine hemoplasmas, such as China (Fu et al., 2017), South Korea (Seo et al., 2019), and Germany (Stadler et al., 2020), have similar clinical signs to those were formerly concluded for M. suis infection. Porcine hemoplasmas (PHs) have been detected in the biggest three pork producers worldwide [China (Song et al., 2014), the USA (Guimaraes et al., 2011), and Brazil (Sonalio et al., 2020)], as well as Germany (Normand et al., 2020), France (Brissonnier et al., 2020), Japan (Hornok et al., 2018), and Argentina (USDA, 2020).
Avian Infection
Avian mycoplasmosis is caused by four pathogenic mycoplasmas, MG, M. synoviae (MS), M. meleagradis (MM), and M. iowae (MI). The MG and MS are OIE-listed respiratory pathogens that have been causing huge economic losses due to their dramatic drop in egg production, hatchability, weight gain, and feed conversion efficiency. On the other hand, they increase embryo mortality, carcass condemnation, and prophylaxis and treatment costs in layers, broilers, and breeders flocks (Yadav et al., 2021). MG is a major Mycoplasma affecting poultry; it causes symptomatic as well as asymptomatic infections. Clinically, it causes chronic respiratory disease in chickens with difficult breathing, sinusitis, airsacculitis, increase embryo mortality in layer parents, and reduce carcass quality in broilers. More seriously, asymptomatic infection also has a formidable impact on the birds as it can be a predisposing factor to more severe secondary bacterial infections. In addition, MG may predispose the animal to many viral contagious diseases such as Newcastle disease and infectious bronchitis (Michiels et al., 2016).
More importantly, MG has been isolated from many different bird species acting as reservoirs for commercial poultry. For example, it was identified in the tracheal swabs of racing pigeons. However, the examined birds showed unapparent symptoms, they could play a role as the potential carriers of the organism (Tsai and Lee, 2006; Michiels et al., 2016). House finches and other passerines, another free-flying avian species, are regarded as the most serious threat for uncontrollable MG infection transmission. Luttrell et al have conducted a field survey for the assessment of MG prevalence among these bird species. The testing indicated that 19.1% of 671 birds caught at farms and 11.6% of 387 birds caught at feeder sites had a positive result (Luttrell et al., 2001). MS infection sometimes remains asymptomatic, otherwise, it can show signs of lameness, synovitis, mild lower respiratory signs, and airsacculitis (Yadav et al., 2021).
MG infection can be transmitted through horizontal and vertical routes, and so prevention and control measures are mainly through biosecurity and vaccination. Live attenuated and/or recombinant live poxvirus vaccines are commercially available against MG and MS infection. Also, avirulent MG live strains (F, ts-11, and 6/85 strains) can be used safely (Yadav et al., 2021). New research proved that 3 consecutive doses of MG vaccines, one live followed by two inactivated vaccine doses, provide good protection in layers (Kiers, 2020). In terms of advantages and limitations of MG vaccines, the F strain, a field strain with moderate virulence, is preferable in places where wild-type MG is highly virulent because it can defeat this virulent MG strain. The other MG vaccines, ts-11 and 6/85, were used more safely because they were less pathogenic and transmissible toward young progeny. While, they showed a lower potency in field challenge than F strain (US Animal Health Association, 2006). The MG 6/85 vaccine strain was developed through serial passages of a field isolate originating from the United States On the other hand, MG bacterins are becoming less popular in commercial flocks, where long-term control of MG infection is critical issue. Further, bacterins are more expensive and inappropriate as they need individual vaccination of birds (Ishfaq et al., 2020).
The ts-11 strain of MG and MS-H strain of MS are temperature-sensitive strains; both of them were proved to be safe and effective for protection against challenge in both chickens and turkeys when administered by eye drop. Globally, both of them are commercially available. For instance, its administration in Australia has greatly reduced the prevalence of disease in chickens causing a tenfold reduction in the use of macrolides in poultry (Browning et al., 2011).
The strain ts-11 vaccine, a mutant induced by chemical mutagenesis, can produce long-term immunity in chickens, but the protective immunity obtained by this vaccine is dose-dependent. A strain ts-304 has been isolated from ts-11 and demonstrated to be as safe as the ts-11 strain. Surprisingly, it also has been effective but at a lower dose and protective against challenge with the MG wild-type strain. In addition, its protection lasts for at least 57 weeks after a single vaccination at 3 weeks of age (Kulappu Arachchige et al., 2021). Since live vaccines are used in many parts of the world, Sulyok and his team have developed new highly specific molecular methods to rapidly differentiate MG vaccine strains from field virulent isolates using clinical samples (Sulyok et al., 2019).
MI is primarily infecting turkeys and occasionally chickens. The natural MI infection in turkeys results in late embryo mortality, a drop in hatchability, and leg abnormalities in young chicks (Pritchard and Balish, 2015). MM is responsible for air sac disease and musculoskeletal and reproductive disorders mainly in turkeys, it also has been isolated from chickens (Béjaoui Khiari et al., 2011).
Equine Infection
Mycoplasma infection was rarely reported in horses; however, M. felis has been isolated from pleuritis and lower respiratory tract infection cases in equines (Wood et al., 1997). In Japan, using genomic DNA for nanopore sequencing, M. felis strain Myco-2 has been detected from a tracheal wash sample of a diseased horse that suffered from respiratory manifestations. This strain has 98.2% identical nucleotides to the typical reference feline strain ATCC 23391 (Kinoshita et al., 2020). In addition, M. equigenitalium (equi) is a potential cause of infertility, endometritis, and abortion in mares, besides, reduced fertility in stallions (Tortschanoff et al., 2005). It is difficult and time-consuming to identify M. equi in clinical samples, and thus, Nehra et al., have developed a species-specific PCR for M. equi diagnosis in clinical samples (Nehra et al., 2015). Two unidentified Mycoplasma strains (N3 and NI1) isolated from the equine respiratory tract were proven to have cross-reactions with strains of Mmm and M. mycoides subsp. capri (Mmc) (Lemcke et al., 1981).
M. equirhinis was isolated from 10.2% of tracheal wash samples from racehorses in Great Britain (Cardwell et al., 2013) and 16.2% from thoroughbred horses in Turkey (Mete and ÖZGÜR, 2017). More recently, using the loop-mediated isothermal amplification (LAMP) assay, M. equirhinis was isolated from 40.0% of Japanese horses (Uchida-Fujii et al., 2021).
Equine hemoplasmas were discovered for the first time in Germany in 2010, as a new species of hemoplasma (Candidatus M. haemobos- like species). After that, scientists have recorded their incidence (26.5%) using a novel real-time PCR assay (Dieckmann et al., 2012). The chronically infected animals could act as reservoirs of infection to other in-contact animals; a recent study discovered M. ovis-like species in an index horse case. What’s more amazing is that the molecular and phylogenetic analysis of the haemoplasma sequences had 100% identity with 16S rRNA of M. ovis, a hemoplasma mainly related to sheep and goats (Kalantari et al., 2020). The previous discovery means that interspecies transmission of Mycoplasma infection could occur anytime. Based on the results of R segment analysis, a species of human Mycoplasma is a group of strains that share R-segments with average nucleotide identity (ANIs) ≥97%. Moreover, R-segments are superior to 16S rRNA gene sequences and multilocus sequences for the identification and phylogenetic analysis of human Mycoplasma species and their strains (Roachford et al., 2019).
HM infection in horses seems to behave subclinically with low bacterial blood loads as represented by Dieckmann et al. (2012). In addition, the infected horses can act as a potential reservoir of infection by M. ovis-like species for both sheep and humans (Kalantari et al., 2020). Further, Manguin et al. have screened the tracheal microbial inhabitants in asthmatic horses with qPCR and determined that Mycoplasma spp. were included in the microbiome composition of tracheal mucus in horses and asthmatic children, as well (Manguin et al., 2020).
Canine Infection
More than fifteen different Mycoplasma species have been isolated from dogs. They are mostly commensal organisms with a few harmful agents. M. cynos was significantly associated with lower respiratory tract (LRT) disease in dogs. On the other hand, no significant association was detected between M. canis, M. spumans, and M. edwardii and clinical signs of canine LRT disease (Jambhekar et al., 2019). M. spumans and M. maculosum were identified by PCR and sequencing to be responsible for fertility problems in male and female dogs (Tamiozzo, 2021).
M. hemocanis (Mhc) and Candidatus M. haematoparvum (CMhp) are two hemoplasmas species that have been reported in canines. In Italy, symptomatic infection by CMhp in a dog was firstly reported by Rosanna et al., who recommended PCR as a gold standard technique for clinical diagnosis of this pathogen (Rosanna et al., 2020). In Korea, the index case of Mhc infection was reported in a dog showing clinical signs of severe hemolytic anemia (Kim et al., 2020).
M. cynos causes upper respiratory disease in dogs, and it is proved to be associated with increased severity of canine respiratory disease complex (CRDC). Clinical signs may include cough and accumulation of mucus and exudate. Potentially, this microbe often evades the immune response predisposing animals to chronic and secondary bacterial infections (Chalker, 2005).
Feline Infection
Four species of feline hemoplasmas have been characterized in domesticated cats. They include M. haemofelis (Mhf), Candidatus M. haematoparvum-like, Candidatus M. haemominutum (CMhm), and Candidatus M. turicensis (CMt). In China, the first identified feline hemoplasma in cats was Candidatus M. turicensis (CMt) (Zhang et al., 2021b). In Thailand, another study estimated that 16.1%, 24.5%, and 1.6% of the random samples collected from stray cats were infected with Mhf, CMhm, and CMt, respectively (Kamyingkird et al., 2021). In Russia, the estimated prevalence of CMhm, Mhf, and CMt was 7.6%, 5.5%, and 0.7%, respectively (Demkin and Kazakov, 2021). CMhm is the most common type of Mycoplasma species producing hemolytic anemia. Mhf causes a more severe and fatal form of hemolytic anemia in cats, whereas CMhm and CMt have lower severity, but only cause severe infection in immunocompromised cats (Willi et al., 2006). Feline infectious anemia is a disease condition of cats accompanied by severe anemia upon erythrocyte disruption. It is induced following infestation by infectious agents such as hemoplasmas (previously mentioned) and Bartonella species (intracellular vector-transmitted pathogens infecting cats) (Zhang et al., 2021b). The non-hemotropic Mycoplasma (M. felis) causes different disease lesions in cats including conjunctivitis, respiratory symptoms, and polyarthritis (Greene and Chalker, 2012).
Wild Animal Infections
For tortoises, Origgi and Jacobson stated that the most significant bacterial disease that seriously affects the endangered free-ranging and captive tortoises is mycoplasmosis (Origgi and Jacobson, 2000). Mycoplasmas cause upper respiratory tract infection in threatened species, including gopher and desert tortoises in the USA. More specifically, The M. alligatoris causes pneumonia, synovitis, and polyserositis in American alligators (Valentine-King et al., 2020). CCPP affects various species of ungulates, chiefly wildlife species, such as gazelles and some species of antelope-like gerenuks (Baziki et al., 2020). M. ovipneumoniae, another Mycoplasma species, causes pneumonia in wild caprines (Jaÿ et al., 2020). This pathogen possesses LAMPs that are considered to be the most potent stimulator of inflammatory cascades (Bai et al., 2020).
More disturbingly, many recent studies have emphasized the importance of cervids as reservoirs for mycoplasmas infection since these species are considered the essential food source for many predators (Petri et al., 2020). Boes et al. have demonstrated the first natural HM infection in white-tailed deer. Following the high identity of 16S rRNA to the previously described M. ovis organism (Messick et al., 1998), the hemoplasma detected in his study likely represents a strain variation of M. ovis, an erythrocytic parasite of ovines (Boes et al., 2012). André et al. have molecularly detected HM in wild canids for the first time in Brazil that are regarded as endangered species; therefore studies concerning their pathogenic threats to their health are critically concerned (André et al., 2011). M. ovis is a zoonotic pathogen that has already been demonstrated in reindeer (Stoffregen et al., 2006) and white-tailed deer species (Maggi et al., 2013a) in captivity in the USA as well as in free-ranging spotted deer species in Japan (Watanabe et al., 2010). In Brazil, M. ovis has been detected in free-ranging marsh deer and pampas deer species (Grazziotin et al., 2011). A recent study identified for the first time, the occurrence of M. ovis in the gray brocket deer and small red brocket deer in the Brazilian national conservation plan for endangered South American deer (André et al., 2020).
M. conjunctivae, an important contagion of wild caprinae, cause infectious keratoconjunctivitis (IKC) in the form of mild symptoms in domestic sheep and goats, while it provokes a severe inflammation of conjunctivae and cornea in wild caprinae. It was responsible for severe epidemics episodes in wild caprinae including chamois and ibex (Marco et al., 2009). In the most advanced stages of IKC, corneal ulceration and perforation, as well as 30% mortalities, have been reported. Eye blindness is a consequence of bilateral eye infection which increases the fatality rate, especially in steep rocky areas because of the falling of affected animals from cliffs (Giacometti et al., 2002).
In view of the increasing transmission of MG to house finches in the wild, and alarmingly, it was responsible for the death of over 200 million birds (Nolan et al., 1998), this marked the first epidemic of MG in the wild birds. Besides, the excessive speed at which this pathogen goes rampant among the house finch population illustrates the rapid pathogen dissemination throughout a large geographic area within a very gregarious and mobile host population (Fischer et al., 1997). Till now, MG has been expanding its host range. For instance, it was identified in many phylogenetically different birds including songbirds (Fischer et al., 1997), raptors (Wrobel et al., 2016), and wild passerines (Luttrell et al., 2001). Hence, rapid evolutionary changes of the pathogen as it expanded geographically allow it to be one of the most recognized wildlife pathogen outbreaks (Sawicka et al., 2020). A possible explanation of the aforementioned host diversity or switching might be due to a shift in CRISPR system dynamics. Also, the gradual degradation and critical functional loss of the CRISPR system in house finches MG after the host switch appears to have a great impact on the pathogen evolution (Delaney et al., 2012). More recently, Mycoplasma infections have been found in migratory wild geese, while, the question concerning the pathogens’ transmission and dispersion is still poorly understood (Sawicka-Durkalec et al., 2022)
Laboratory Animals
Laboratory Animals’ Infection
Laboratory animals are useful fundamental scientific tools; the progression of apparent or in-apparent infections with Mycoplasmas has tremendous alterations on the normal physiological responses of mice throughout experiments. When mycoplasmas are running rampant in the experimental animals’ population, the only solution will be through introducing specified pathogen-free (SPF) animals and animal facilities to avoid potential false results. M. collis, M. pulmonis, M. neurolyticum, M. muris, and M. arthritidis are the most common Mycoplasma spp. infecting mice (Masoumalinejad et al., 2018).
M. pulmonis is the most prevalent Mycoplasma pathogen in mice causing otitis media, reproductive disorders, as well as substantial respiratory consequences with a prevalence of (20-60) % (Booth et al., 2014). The most important issue regarding M. pulmonis infection in rodents is that it is probably the best model from which we have learned the most about the determinants of immunity to control human M. pneumoniae respiratory infections. Many studies have used the experimental M. pulmonis rat infection model as an ideal model in terms of the ciliary cell function and cellular kinetics (Lambert et al., 1998), neurogenic inflammation (McDonald et al., 1991), natural killer cell activity (Kamiyama et al., 1991), local and systemic immune response (Steffen and Ebersole, 1992), induction of the production of several cytokines (Faulkner et al., 1995), and polyclonal proliferation of B and T lymphocytes (Rocha Sobrinho et al., 2011). M. neurolyticum has been characterized as a mammalian brain organism responsible for nerve disorders as a result of secreted Mycoplasma toxins (Tully, 1981). Also, M. collis was isolated for the first time from the conjunctiva and nasal cavity of mice and rats (HILL, 1983). M. muris, a scarce Mycoplasma type causing a huge hazardous effect on the reproductive efficiency of female mice, has been identified in recent years (Zinatizadeh et al., 2017). M. arthritidis, another rare pathogen of mice, is regarded as the main cause of arthritis in mice with swelling of legs and fingers (Constantopoulos and McGarrity, 1987).
Animal Models for <italic>Mycoplasma</italic> Infection
The laboratory mice are the most common species used in animal experimentation in biomedical research. In addition, the experimental mouse mastitis model allows us to examine a large number of Mycoplasma strains (Dmochowski, 1967).
Saraya et al., have designed five mouse models for M. pneumoniae pneumonia to examine the pathological picture in animals with various immune statuses. Firstly, animals were immunized following different regimes (one for each animal model). Afterward, they were challenged with M. pneumoniae antigen intratracheally, only mice groups immunized with M. pneumoniae antigen and alum adjuvant or M. pneumoniae antigen with CpG adjuvant (Th2 predominant) have developed severe lymphoplasmacytic infiltration in the peri-bronchovascular areas (PBVAs). These results indicate that the adaptive host immune responses in these two models seem to be the main regulator for human M. pneumoniae pneumonia pathological features and Th2 predominant characteristics might be important to generate and simulate the typical picture of M. pneumoniae pneumonia (Saraya et al., 2014).
In vivo strategies whereby BALB/c mice were injected subcutaneously with the T-B epitope peptides resulted in strong antigen-specific serum antibody and cellular immune responses, besides decreasing the inflammatory response of the challenged mice with M. pneumoniae (Weng et al., 2017), we can take the advantage of these findings for Mycoplasma vaccine production.
Several examples of useful Mycoplasma animal models including gerbils (burrowing mouse-like rodents) inoculated intranasally with M. pneumoniae to investigate its pathogenesis of human lung infection (Rodríguez et al., 2021). A rabbit model was used in previous studies for the development of polyclonal and monoclonal antibodies against various human diseases (Kaur et al., 1998). Furthermore, hamsters were injected intratracheally with M. fermentans culture to explain the ability of the pathogen to induce pneumonia and chronic infectious diseases in humans (Yáñez et al., 2013).
Guinea pigs are regarded as the best animal model after non-human primates to study M. pneumoniae infections. In an important study, Dumke et al. have used these animals for studying the pathogen-host relationship as well as characterization and subtyping of M. pneumoniae strains isolated from human patients. The adaptation, preference, and survival of individual strains also have been investigated. They concluded that M. pneumoniae species is genetically highly homogeneous (Dumke et al., 2004). Hausner and his team have also immunized guinea pigs with a hybrid protein composed of adherence-related parts of the proteins P1 and P30 of M. pneumoniae. The results showed a dramatic decrease in its detection in pulmonary samples from vaccinated as well as subsequently infected animals (Hausner et al., 2013). Meanwhile, sera from immunized animals have been demonstrated to have crucial adherence-blocking properties. Besides, the initiation of potent stimulation of mucosal immunity was the milestone for successful vaccination with intranasal antigen as well as in combination with other biocompatible adjuvants (Zhu et al., 2012).
In vivo studies using non-human primates have also played a vital role to investigate the antigenic and immunogenic properties as well as the pathogenicity of specific mycoplasmas including M. genitalium. For instance, experimentally infected primates have been used to examine M. genitalium membrane topology, antibody accessibility, amino acid diversity, and the location of functional and antigenic epitopes for the MgpB adhesion (Iverson-Cabral et al., 2015). Another important example of using primates instead of humans for doing essential experimental work is macaque (a genus of Asian monkeys) which was used for studying the persistence, immune response, and antigenic variation of M. genitalium in an animal experimental infection model (Wood et al., 2013).
Large animal models were also established for the Mycoplasma study. The experimental infection of SPF lambs with M. ovipneumoniae resulted in the establishment of asymptomatically infected upper airways in absence of other secondary infections (Davies et al., 1981). M. bovis calf infection model was used to estimate the effectiveness of some antimicrobial agents against animal mycoplasmas (Dudek et al., 2019). These previously illustrated experimental models to study mycoplasmas are valuable in many points of view such as investigation of their infection pathogenesis and immune response, assisting in the development of therapeutic strategies and diagnostic biomarkers, and conducting the potential vaccine candidate’s trials. On the other side, some encountered limitations should be taken into account as the availability of SPF conditions to avoid false results.