Analysis of the Indigenous and Pathogenic Microbiota in Bone Sequestration due to Osteonecrosis of the jaws by Bisphosphonates (MRONJ)

Bisphosphonates (BP) are a pharmacological class of synthetic inorganic pyrophosphate analogs that have an affinity for calcium. They are used in the treatment of various benign and malignant metabolic conditions, such as Paget's disease; multiple myeloma; and metastases from distant sites such as breast, thyroid, prostate, and lung. The oral form of BP is indicated in the treatment of osteoporosis, fibrous dysplasia and, more recently, imperfect osteogenesis in the pediatric population. 

The American Surgery of Bone Mineral Research (ASBMR) defined MRONJ as “necrotic bone area exposed to the oral environment with more than eight weeks of permanence, in the presence of chronic treatment with BPs, in the absence of radiation therapy to the head and neck”in 2007 [1]. 

American Association of Oral and Maxillofacial Surgeons (AAOMS) in 2014 divided the MRONJ into 4 stages from 0 to 3 according to the clinical and radiological aspect of the osteonecrotic lesion: stage 0: Osteonecrotic lesion without sign-pathognomonic 1: osteonecrotic lesion with clinical signs and but no clinical symptoms; Stage 2: Osteonecrotic lesion with sign and clinical symptoms; Stage 3: Osteonecrotic lesion with signs and symptoms that involve other structures such as: pathological fractures, anesthesia of the lower dental nerve, oral-nasal communication, oral-sinus communication, skin fistulas [2]. 

Bone exposed for at least 8 weeks in duration with or without pain it can signify the diagnosis of MRONJ. In studies of exposed bone, colonization of Actinomyces spp. without knowing if they play a critical role or is a secondary infection in the pathogenesis of MRONJ. Actinomyces are part of the normal microbiota of the oral cavity, however, it is difficult to determine whether it represents microbial colonization in hard and soft tissues or is a true infection, being the clinical experience, that this pathogen Microbial is an opportunistic bacterium directly involved in the pathology of the hard and soft tissues of the maxillofacial region and oral cavity in the MRONJ patient [3-5]. 

Ruggiero, et al. evidenced any cases in which the diagnosis of Actinomyces was established, bone exposure was not always observed, which corresponds to incipient stages in the exposure document of the American Association of Oral and Maxillofacial Surgeons [6]. A review published cases identified multiple bacterial species in patients with MRONJ, the main one being Actinomyces [4,7], but other microbial pathogens are also evident, such as: Streptococcus, Staphylococcus, Treponemes, Bacteroides, Actinobacillus, Moraxella and Eikenellacorrodens [5,8]. It is believed that these microorganisms exert a synergistic effect on the pathogenesis of the disease, secreting bacterial enzymes, such as collagenases and hyaluronidases, which destroy tissues and promote the extension of the lesion to other areas of the head and neck. 

It is known that the microbiological culture in an anaerobic medium will be carried out when empirical antibiotic therapy is not effective, a decision made together with the treating Antibiograms are generally not applicable to determine the correct treatment of MRONJ since the bacterial flora found corresponds to the usual pathogenic oral microbiota: Phorphyromona gingivalis and Aggregatibacter actinomycetemcomitans [9]. Therefore, based on the scientific evidence, it is currently not known whether infection of the jaws and overlying soft tissues is a primary or secondary event in MRONJ [9-11]. Management of MRONJ remains a significant clinical challenge. The objective of this article is to explore current theories about the importance of infection in the development of this devastating pathology.

There is clinical evidence that Actinomyces may play a fundamental role in the pathogenesis of MRONJ associated with bisphosphonates: antiresorptive and antiangiogenic. Identification and a prolonged course of oral antimicrobial therapy can lead to complete resolution of this actinomycotic osteonecrosis. Failure to identify Actinomyces spp. and initiate antibiotic treatment could be responsible in many cases for making treatment difficult and refractory [12-18]. The detection of Actinomyces spp. in 89% of bone samples by histology is remarkable, but there may still be a significant underestimation of the true frequency of MRONJ associated with this infection [19]. Some authors recommend beta-lactam agents since they have a high therapeutic index that allows the safe administration of high doses of drugs and high levels of therapeutic drugs are reached in serum, tissues, bile and synovial fluid. 

Steininger, et al. stated that Actinomyces spp. isolates are universally susceptible to beta-lactam antimicrobial agents [20] and that treatment without a beta-lactamase inhibitor also significantly reduces the rate of gastrointestinal adverse events such as abdominal discomfort or diarrhea from any cause, including Clostridium difficile enterocolitis. Consequently, they recommend for the treatment of actinomycosis in concordance with others the use of beta-lactam antimicrobial agents in high daily doses before the final surgical treatment of MRONJ. 

For patients with a penicillin allergy, tetracyclines are a good alternative for oral therapy, especially in milder presentations of the disease. In severe infections, carbapenems or the new compound tetracyclines may be appropriate therapeutic options [19-21]. 

Ivaniushko, et al. have identified 15 species of microorganisms demonstrating an important role of oral cavity pathogens in the development of osteonecrosis of the facial bones. Enterobacteriaceae were identified; Streptococcus spp; Fusobacteriumspp; Staphylococcus spp; Prevotella; Porphyromonas; Megasphera; Veilonella; Dialister; Corynebacterium; Mobiluncus; Leptotrichia; Sneatia; Lachnobacterium; Clostridium; Peptostreptococcusspp; ?ubacteriumspp; Candida spp in maxillary osteonecrosis of patients [22]. It is believed that the exposure of bone to the oral environment generates the appropriate conditions for the microbial colonization of all these species, and that they act by aggravating the clinical and exacerbate symptoms in the area. 

Based on comparison above, other parts of the body, bone can be easily colonized by the abundant microbiota of bacterias and yeasts in the oral cavity (including periodontal pockets and periapical abscesses). Various studies confirm that the use of systemic antibiotics cannot restrict the colonization of bacteria after the initiation of MRONJ [23]. There is no consensus on how is the best way patient´s be prescribed antiresorptive medication. Our protocol is exhaustive with the use of antiseptics to control the oral microbiota, so that it does not colonize bone sequestration and diffuse other spaces due to contiguity or continuity. Studies have demonstrated that a onetime use of chlorine dioxide containing mouth rinse significantly improves mouth odor pleasantness, reduces mouth odor intensity, and reduces volatile sulfur compound concentrations in mouth air for at least 8 hours after use [24]. 

Chlorhexidine is widely used in mouthwash solutions, but is also found in dental gels and toothpaste. Chlorhexidine in mouthwash solutions binds to oral mucosal surfaces via electrostatic forces, inhibits dental plaque formation and exerts a bacteriostatic action that persists for several hours [25]. Povidone-iodine mouthwash is the only approved mouthwash for pre-procedural rinsing in dental practice as antibacterial, antiviral, and antifungal effects according to the available literature [26]. At last, Cetylpyridinium chloride is the most common and effective mouthwashes used in dental practice [27]. 

It is essential patients with MRONJ be treated in an interdisciplinary fashion. Stomatognathic system should be examined preventatively prior to the initiation of BP, DS or anti angiogenic drugs treatment in order to avoid pathological buccal manifestations, following the same healthcare clinical protocols are used for patients who receive head and neck radiotherapy. Additionally, patients should be informed of the precautions to be taken, including regular dental appointments for oral health assessment. The risk of developing MRONJ should be evaluated according to the type of AR or anti angiogenic drugs which are indicated depends on treatment duration. [27, 28].

MRONJ Etiopathogenesis has not been fully elucidated, although there are many theories that attempt to explain it. Some authors point in infection as the main cause and not just a secondary event to bone exposure [29,30]. Others authors publicated that bone impregnated with AR is less resistant to bacterial infection and there is colonization by the normal microbiota as an ideal incubator for periapical periodontal microorganisms, which stimulate chronic inflammation [31]. Also other authors, who suggest an electrostatic interaction between nitrogen-containing bisphosphonates and Gram-positive bacteria, which would favor bone infection by these microorganisms [32]. 

The main colonizer is Actinomyces spp, but other oral microorganisms (including Candida spp) can be favored by the metabolic conditions, mechanical retention and microbial succession to lodge in necrotic bone. Therefore, based on our protocols, the use of antiseptics is highly recommended to control colonization with microorganisms avoid oral microbiota. Recommending the use of Chlorhexidine on a rotating basis; Povidone Iodine; Cetylpyridinium Chloride and Rifamycin to avoid generating microbial resistance to them, Antiseptic washes were started with Chlorhexidine 0.12%, Povidone Iodo 10%, Rifamycin 0.05% and Cetylpyridinium Chloride 0.5%alternating them fifteen days each other, in order to produce the reflux of the inflammatory content, opportunely accompanied by antibiotic therapy: in the first instance amoxicillin 500mg with clavulanic acid 125 mg every 8 hours, in case of a lack in the recovery response at 72 hours, metronidazole 500 mg is added. In severe MRONJ situations we prefer to use ciprofloxacin 500 mg every 12 hours for 7 days [27]. Prophylactic antibiotic is not necessary to clinical attention depend on MRONJ so, patients with MRONJ have no risk of bacterial Endocarditis, except patients like oncology comorbidity requires [28, 33,34].