Coronavirus disease 2019 (COVID-19), caused by a severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has caused a global pandemic and resulting serious public health crisis (Zhu et al., 2020; World Health Organization, 2022). While most COVID-19 patients have minor symptoms, ~15% of hospitalized patients require admission to an intensive care unit (ICU; Terlecki et al., 2021). These patients exhibit respiratory failure with a systemic inflammatory reaction and multiple-organ dysfunction, requiring oxygen supplementation and, in some cases mechanical ventilation (Weiss and Murdoch, 2020).

Over 1,000 bacterial species have been reported to reside in the oral cavity (Dewhirst et al., 2010). In healthy individuals the oral bacteriota is dominated by Actinobacteria (Acctinomyces, Corynebacterium and Rothia), Bacteroides (Capnocytophaga, Porphyromonas and Prevotella), Firmicutes (Granulicatella and Streptococcus), Fusobacteria, Proteobacteria (Haemophilus and Neisseria; Zaura et al., 2001). The oral cavity can be divided into several microbiologically distinct niches, including saliva, soft tissue surfaces of the oral mucosa and tongue, and hard tissue surfaces of teeth (Xu et al., 2015; Zhang et al., 2018). Of the most common species, 54% is cultivable and identifiable, 14% is cultivable but not easily identified, and 32% cannot be cultivated, remaining in dormant state (Caselli et al., 2020). Recent research of oral microbiota resulted in two large databases: Human Microbiome Project (HMP) and Human Oral Microbiome Database (HOMD). HMP contains microbiome data from 5 main environments: the oral cavity, nasal cavity, vagina, gut and skin. Data in HOMD encompasses oral microbiota composition (Li et al., 2022).

Previous studies of changes in microbiome, including the oral microbiota, showed a significant reduction of microbial diversity in SARS-CoV-2 affected patients (Iebba et al., 2021; Wu et al., 2021; Uehara et al., 2022). These changes comprised decreased abundance of Neisseria, Corynebacterium, Aggregatibacter, Treponema, and Pseudomonas genus, and Prevotella intermedia in the oral cavity of COVID-19 patients. Importantly, the loss of comensal Neisseria, such as N. subflava and N. mucosa, and Prevotella spp. acting as a local oral probiotic, can lead to severe imbalance in the oral microbiota composition (Weyand, 2017; Rafiqul Islam et al., 2022). Enrichment of Campylobacter, Granulicatella, Veillonella and Filifactor genus was also observed, that can be of clinical significance, as those are taxa associated with periodontitis (Wu et al., 2021). Veilonella spp. has been also reported to induce proinflammatory responses (Haran et al., 2021). Furthermore, increased abundance of opportunistic Hemophilus parainfluenzae in the oral cavity can predispose patients to respiratory tract infections (Iebba et al., 2021; Wu et al., 2021). Another report revealed that COVID-19 patients had a higher abundance of Enterococcus spp. in the oral cavity, linking respiratory pathogens with gut microbiome abnormalities (Rafiqul Islam et al., 2022). The dysbiosis was even more pronounced in severe course of infection and long-COVID-19, suggesting that its extent can be treated as an indicator of infection severity (Haran et al., 2021; Wu et al., 2021; Fiema et al., 2022; Rafiqul Islam et al., 2022). The use of antibiotics in COVID-19 patients was also associated with independent oral and gut microbiome profiles (Wu et al., 2021).

A high prevalence of oral health problems, such as xerostomia, mucosal blistering, mouth rash and lip necrosis has been observed in patients with COVID-19 (Aragoneses et al., 2021). Several trials have correlated poor oral hygiene with hyper-inflammation (Kamel et al., 2021), and poor oral health in patients with caries and periodontitis may play a significant role in the development of severe complications of COVID-19 in patients managed in the ICU (Hocková et al., 2021; Marouf et al., 2021). Moreover, during prolonged endotracheal intubation, dysbiotic oral microbiota can colonize the lower respiratory tract. These patients are at high risk for developing bacterial ventilator-associated pneumonia (VAP). The oral management of these patients in an ICU is critical as oral care has been shown to reduce the incidence of VAP (Bao et al., 2020; Luyt et al., 2020).

The aim of this study was to evaluate the oral health status and assess the dysbiosis of cultivable oral bacteriota in COVID-19 patients hospitalized in an ICU with acute respiratory distress in the early post-intubation period.

In this report, we comprehensively characterized the oral health condition and cultivable oral bacteriota in COVID-19 patients hospitalized in an ICU with ARDS within 36 h following intubation. In this population, the oral microbiota from mucosal swabs, dental samples, and gingival pockets showed significant qualitative and quantitative dysbiosis and was distinct from healthy patients. SARS-CoV-2 infection, hospitalization in an ICU and mechanical ventilation are important factors leading to oral dysbiosis in patients.

Our study population comprised a homogenous group of patients with COVID-19 infection that were hospitalized in a temporary ICU dedicated to SARS-CoV-2-positive patients. These patients, with severe COVID-19 and ARDS, required specialist medical care, including mechanical ventilation or hemodialysis. Our data demonstrated a significantly higher mortality rate compared to normal COVID admissions, although our election criteria biased patient selection toward the most severe COVID-19 cases. Our study population was also older, with multiple comorbidities that have confirmed deleterious effects on COVID-19 survival, such as diabetes and cardiovascular disease (Harrison et al., 2021).

Our study population presented with moderate or severe dysfunction of oral health. We believe that this resulted from first, poor oral health status in the elderly adult Polish population as presented in one recent report (Malicka et al., 2022). The authors noted that 21% of 70-year-olds were completely toothless. On average, the number of teeth was 12.97 ± 9.5., with 4.7 ± 4.8 teeth in the occlusion. 30.8% of patients wore a partial removable denture, and 25% a complete removable maxillary denture. 20% of study participants had a partial removable denture, and 22.6% had a complete removable mandibular denture. Oral dryness was observed in ca. one-third of the studied cohort, nearly 20% had periodontitis and ~30% required treatment for caries. They emphasized that more than 60% of patients required professional dental prophylaxis (Malicka et al., 2022). Secondly, another report from a similar cohort (age and comorbidities) revealed patients hospitalized due to myocardial infarction presented higher plaque and periodontal indices as compared to patients with stable angina pectoris (Wozakowska-Kapłon et al., 2013). Still, this subject is under researched and further exploration is required. Finally, previous studies of ICU COVID-19 patients reported xerostomia, mucosal blistering and ulcers, rash, lip necrosis, and loss of taste and smell (Kamel et al., 2021; Eduardo et al., 2022; Yoshino et al., 2022). These can exacerbate such conditions as periodontitis, being an important risk factor for complications in patients with COVID-19 hospitalized in the ICU (Marouf et al., 2021). Pre-COVID-19 reports showed that hospitalization in an ICU together with mechanical ventilation can have a deleterious effect on oral health (Terezakis et al., 2011), including accumulation of the dental plaque and emergence of mucosal lesions. Due to the deterioration of oral health, critically ill patients in the ICU represent a group vulnerable to further complicates including VAP (Luyt et al., 2020).

Another noteworthy finding was a significant and alarming qualitative and quantitative dysbiosis of the cultivable oral bacteriota in our study population, as early as up to 36 h following intubation. Potentially pathogenic bacteria including Acinetobacter baumannii, Enterococcus faecalis, Escherichia coli and Klebsiella pneumoniae occurred frequently with large CFU counts in our study population. While previous studies showed that these species are found in the oral cavity, their prevalence was not as high as in our patient population (Beck, 1979; Marshall et al., 2020; Diagnostic testing for SARS-CoV-2 [internet], 2022).

The CFU counts of such species as Enterococcus faecalis and Acinetobacter baumannii were as high as commensal Streptococci and Staphylococci, indicating significant abnormalities in the oral bacteriobiota hemeostasis.

In one study, oral rinse samples from COVID-19 patients with a wide spectrum of symptoms showed a comparable extent of oral dysbiosis, with lower bacterial diversity, higher abundance of Lactobacillus spp., Enterococcus spp., Acinetobacter baumannii, and lower amounts of Gemella spp., Fusobacterium spp. and Haemophilus spp. (Soffritti et al., 2021).

The prevalence of E. coli was surprisingly high in our population. Escherichia coli is not a member of commensal oral microbiota, however it was detected in oral cavities of elderly patients with systemic diseases (Zawadzki et al., 2017). Recently, E. coli was reported to successfully colonize a supragingival biofilm (Pérez-Chaparro et al., 2014), so under special nutritional and environmental circumstances, E. coli can likely survive and even dominate this niche, especially in immunocompromised patients (Thurnheer and Belibasakis, 2014). Additionally, we found a higher prevalence of E. coli in patients admitted from the emergency ward than in those transferred from other hospital wards. To date, there have been no other reports on this issue, warranting further research.

Conversely, Enterococcus faecalis was more frequent in patients transferred from other hospital wards than those originating from the emergency department. Previous studies showed similar results, with changes in the oral bacteriobiota composition result from the exposure to hospital bacteria and each subsequent day of hospitalization increases the risk of Enterococci infections (Russo Fiorino et al., 2021). Moreover, E. coli and E. faecalis can employ antagonistic interactions against S. mutans (Thurnheer and Belibasakis, 2014), partially explaining our observations.

Our analyses revealed that the prevalence of commensal Streptococcus strains was lower in patients with a higher antibiotic consumption prior to intubation. We consider that antibiotics could be a major factor contributing to oral dysbiosis and disappearance of “healthy” commensal strains. Of all antibiotics, β-Lactam antibiotics were most commonly used in our cohort. Prospective cohort studies revealed that Shannon biodiversity index was decreased during amoxicillin treatment and was subject to further reduction in the following 6 months’ time period (Menon et al., 2019; Monroy-Pérez et al., 2020; Nel Van Zyl et al., 2022). The density of Neisseria, Streptococcus and Veillonella strains in the oral cavity was also reported to decrease during treatment with amoxicillin (Larsson Wexell et al., 2016; Moraes et al., 2020). Save commonly used amoxicillin, other groups of antibiotics with various mechanisms of action can influence the oral microbiota and promote the selection of multi-drug resistant strains and their horizontal transmission (Zaura et al., 2015; Moraes et al., 2020). Other factors that can lead to oral dysbiosis include: local and systemic diseases, improper oral hygiene, unbalanced diet, smoking tobacco and immunosuppression (Li et al., 2022).

Lactobacillus was more prevalent in survivors in our study population, but there were no associations with antibiotic use, or with probiotic use according to the care standards in UH wards. One previous study reported the relative abundance of various bacterial genera, including Lactobacillus spp. in COVID-19 patients (Soffritti et al., 2021). These findings are notable, as Lactobacillus spp. may play some role in the protection against SARS-CoV-2, acting as an inhibitor of viral contamination by multiple mechanisms, including production of metabolites with antiviral activity, stimulation of mucosal immune system cells and local cytokines production (Zrelli et al., 2021).

Previous studies of COVID-19 patients tested saliva or nasopharyngeal swabs (Lloréns-Rico et al., n.d.; Miller et al., 2021). One notable strength of our study is that we investigated mucosal and dental brushes, highly representative for microbiota analysis (Zaura et al., 2001). Considering a proper and complex oral health evaluation, we used the BOAS scale. Among oral assessment tools, BOAS has been proposed as the most appropriate for ICU patients, with the mucosal-dental plaque score most applicable during observation (Ames et al., 2011).

Our study also had some limitations. First, we focused only on the SARS-CoV-2-positive patients. Moreover, in this study we used traditional methods for identification of microorganisms on the species level. As it is known, NGS it also allows obtaining information on non-cultivable microorganisms. But the method we used allowed the establishment of a microbial bank, for future studies in healthcare-associated-infections.

COVID-19 patients hospitalized in an ICU in the early post-intubation period presented an alarming qualitative and quantitative dysbiosis of the cultivable oral bacteriota. Abnormalities in the oral health status can trigger deterioration and dysbiosis of the oral microbiota. Poor oral hygiene, cough, increased inhalation and mainly mechanical ventilation provide a pathway for oral microorganisms to enter the lower respiratory tract, leading to pneumonia. A proper assessment of oral health can provide information on how to treat and diagnose these patients. Effective oral health care measures are necessary to reduce these infections, especially in severe COVID-19 patients.

The raw data supporting the conclusions of this article will be made available by the authors, without undue reservation.

The studies involving human participants were reviewed and approved by by the Jagiellonian University Bioethics Committee (Komisja Bioetyczna, Uniwersytet Jagielloński). The patients/participants provided their written informed consent to participate in this study.

IG-M: conceptualization, investigation, methodology, project administration, resources, supervision, and writing–review and editing. MF: data curation, methodology, investigation, writing–original draft preparation, and writing–review and editing. MK: formal analysis, methodology, visualization, writing–original draft preparation, writing–review and editing. EJ: formal analysis, methodology, validation. DR: formal analysis, methodology, validation. JW-M: methodology, formal analysis, validation, writing–review and editing, resources. All authors contributed to the article and approved the submitted version.

This publication was supported by the National Center for Research and Development CRACoV-HHS project (Model of multi-specialist hospital and non-hospital care for patients with SARS-CoV-2 infection) through the initiative “Support for specialist hospitals in fighting the spread of SARS-CoV-2 infection and in treating COVID-19” (contract number SZPITALE-JEDNOIMIENNE/18/2020). The described research was implemented by consortium of the University Hospital in Krakow and the Jagiellonian University Medical College.

The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

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