Analyzing the characteristics of respiratory microbiota after the placement of an airway stent for malignant central airway obstruction.

Malignant central airway stenosis is treated with airway stent placement, but post-placement microbial characteristics remain unclear. We studied microbial features in 60 patients post-stent placement, focusing on changes during granulation tissue proliferation. Samples were collected before stent (N = 29), after stent on day 3 (N = 20), and after granulation tissue formation (AS-GTF, N = 43). Metagenomic sequencing showed significant respiratory tract microbiota changes with granulation tissue. The microbiota composition, dominated by Actinobacteria, Firmicutes, and Proteobacteria, was similar among the groups. At the species level, the AS-GTF group exhibited significant differences, with Peptostreptococcus stomatis and Achromobacter xylosoxidans enriched. Analysis based on tracheoesophageal fistula presence identified Tannerella forsythia and Stenotrophomonas maltophilia as the main differential species, enriched in the fistula subgroup. Viral and fungal detection showed Human gammaherpesvirus 4 and Candida albicans as the main species, respectively. These findings highlight microbiota changes after stent placement, potentially associated with granulation tissue proliferation, informing stent placement therapy and anti-infective treatment optimization. IMPORTANCE: Malignant central airway stenosis is a life-threatening condition that can be effectively treated with airway stent placement. However, despite its clinical importance, the microbial characteristics of the respiratory tract following stent insertion remain poorly understood. This study addresses this gap by investigating the microbial features in patients with malignant central airway stenosis after stent placement, with a specific focus on microbial changes during granulation tissue proliferation. The findings reveal significant alterations in the diversity and structure of the respiratory tract microbiota following the placement of malignant central airway stents. Notably, certain bacterial species, including Peptostreptococcus stomatis and Achromobacter xylosoxidans, exhibit distinct patterns in the after-stent granulation tissue formation group. Additionally, the presence of tracheoesophageal fistula further influences the microbial composition. These insights provide valuable references for optimizing stent placement therapy and enhancing clinical anti-infective strategies.

Dysbiosis and Variation in Predicted Functions of the Granulation Tissue Microbiome in HPV Positive and Negative Severe Chronic Periodontitis.

Retrospective analysis has already shown correlation between severe Chronic Periodontitis (CP) cases with human papiloma virus (HPV). Hence, we aimed to explore deep-seated infected granulation tissue removed during periodontal flap surgery procedures for residential bacterial species between HPV+ and HVP- CP cases, which may serve as good predisposition marker for oral cancer. All CP-granulation samples showed the prominence of Firmicutes, Proteobacteria, and Bacteroidetes phyla with an abundance of gram negative anaerobes, except Streptococcus. In Beta diversity nonmetric multidimensional scaling plot, the random distribution of species was observed between HPV+ and HPV- CP granulation-samples. However, an abundance of Capnocytophaga ochracea was observed in HPV+ CP samples (p<0.05), while Porphyromonas endodontalis, Macellibacteroides fermentas, Treponema phagedenis, and Campylobacter rectus species were highly abundant in HPV- CP samples (p<0.05). The differential species richness leads altered functions related to mismatch-repair and nucleotide excision-repair and cytoskeleton-proteins. Hence, differential abundance of gram negative bacterial species between HPV+ and HPV- granulation-samples under anaerobic conditions may release virulence factors which may alter pathways favouring carcinogenesis. Hence, these species may serve as good predisposition marker for oral-cancer.

Novel negative pressure wound therapy device without foam or gauze is effective at -50 mmHg.

Negative pressure wound therapy (NPWT) promotes healing in acute or chronic wounds. Conventional NPWT devices consist of a filler (such as foam or gauze) that covers the wound and of a permeable membrane and tubing that connects the space under the membrane to a suction pump. The permeable membrane increases airflow and thus increases the required pump capacity that can cause patient discomfort or even ischemia in wounds with compromised vascularity. In addition, foam or gauze may fragment and become colonized with bacteria over time. To mitigate these, negative aspects, we have developed a new impermeable single layer component membrane dressing to deliver NPWT that does not need a foam or gauze to function. Therefore, the purpose of this study was to introduce this novel NPWT system (platform wound device, PWD) and evaluate its usability and effectiveness in the treatment of porcine full-thickness burns. A total of 48 burn wounds were created across four Yorkshire pigs on the dorsum. Wounds were created on day 0 and continuous NPWT with -50 mmHg and - 80 mmHg was initiated immediately. Subsequently, the burns were debrided on day 3 and animals were euthanized on day 7. The efficacy of the PWD on wound healing and reduction of bacterial burden was measured and compared to wounds that did not receive NPWT. The results showed that PWD promoted wound healing by outperforming the wounds that did not receive NPWT and that PWD was efficient at reducing bacteria from the burn eschar and from the wound bed. In conclusion, this study demonstrated that PWD promoted wound healing with a negative pressure as low as -50 mmHg, which likely benefits healing and avoids potential safety issues.

Factors impairing cell proliferation in the granulation tissue of pressure ulcers: Impact of bacterial burden.

The authors aimed to assess the factors that impair cell proliferation in the granulation tissue of pressure ulcers using immunohistochemistry for the cell proliferation marker Ki-67. This was a single center, cross-sectional study. The study included 86 patients with stage III or IV pressure ulcers. Two granulation tissue biopsy specimens were obtained from 86 patients. The specimens were used for histological examination, Ki-67 immunohistochemistry, and bacterial count assessment. The % of Ki-67-stained cells was considered as the Ki-67 index. Pearson's product-moment correlation coefficient (r) was used to assess the relationship between the Ki-67 index and other quantitative variables, including age, body mass index, bacterial count (Log10 CFU/g), serum albumin level, hemoglobin level, white blood cell count, and C-reactive protein level. The Mann-Whitney U test was used to compare the mean Ki-67 index according to gender, diabetes, smoking status, and wound culture. Univariate and multivariate linear regression analyses were used to assess the association between the Ki-67 index and other parameters. The Mann-Whitney U test revealed that the bacteria-positive group had a lower Ki-67 index (p = 0.045). Bacterial count demonstrated a significant negative correlation with the Ki-67 index (r = -0.325, p = 0.002). Multivariate linear regression analysis showed that bacterial count was a significant predictor of the Ki-67 index. The adjusted ß-coefficient was -1.34 (95% confidence interval, -2.01 to -0.66, p < 0.001). Among the isolated bacteria, Corynebacterium spp. and Staphylococcus aureus were significantly associated with a low Ki-67 index, but Pseudomonas aeruginosa was not. These results suggest a negative relationship between bacterial count and cell proliferation in pressure ulcer granulation tissue, as indicated by the Ki-67 index. Granulation tissue formation in pressure ulcers may be accelerated if high bacterial load is treated appropriately.

16S rRNA Long-Read Sequencing of the Granulation Tissue from Nonsmokers and Smokers-Severe Chronic Periodontitis Patients.

Smoking has been associated with increased risk of periodontitis. The aim of the present study was to compare the periodontal disease severity among smokers and nonsmokers which may help in better understanding of predisposition to this chronic inflammation mediated diseases. We selected deep-seated infected granulation tissue removed during periodontal flap surgery procedures for identification and differential abundance of residential bacterial species among smokers and nonsmokers through long-read sequencing technology targeting full-length 16S rRNA gene. A total of 8 phyla were identified among which Firmicutes and Bacteroidetes were most dominating. Differential abundance analysis of OTUs through PICRUST showed significant (p>0.05) abundance of Phyla-Fusobacteria (Streptobacillus moniliformis); Phyla-Firmicutes (Streptococcus equi), and Phyla Proteobacteria (Enhydrobacter aerosaccus) in nonsmokers compared to smokers. The differential abundance of oral metagenomes in smokers showed significant enrichment of host genes modulating pathways involving primary immunodeficiency, citrate cycle, streptomycin biosynthesis, vitamin B6 metabolism, butanoate metabolism, glycine, serine, and threonine metabolism pathways. While thiamine metabolism, amino acid metabolism, homologous recombination, epithelial cell signaling, aminoacyl-tRNA biosynthesis, phosphonate/phosphinate metabolism, polycyclic aromatic hydrocarbon degradation, synthesis and degradation of ketone bodies, translation factors, Ascorbate and aldarate metabolism, and DNA replication pathways were significantly enriched in nonsmokers, modulation of these pathways in oral cavities due to differential enrichment of metagenomes in smokers may lead to an increased susceptibility to infections and/or higher formation of DNA adducts, which may increase the risk of carcinogenesis.

Úlcera dolorosa en paciente de raza negra / Painful Ulcer in a Black Man

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Nondermal irritating hyperosmotic nanoemulsions reduce treatment times in a contamination model of wound healing.

Increased microbial burden within the wound often complicates wound healing and may lead to subsequent infection or delayed healing. Here, we investigate a novel topical for addressing wound contamination that utilizes hyperosmotic saccharides with a cell membrane disrupting emulsion. These hyperosmotic nanoemulsions (HNE) were administered topically in a full-thickness biopsy model of wound healing. Results show that HNE were well tolerated in noninfected animals with no indications of dermal irritation or acute toxicity. Additionally, HNE was able to reduce bacterial bioburden (Escherichia coli and Enterococcus faecalis) levels by 3 logs within 24 h when wounds were inoculated with 5 × 10(6) total CFU. These bactericidal values were similar to wounds treated with silver sulfadiazine. Wound closure showed HNE wounds closed in 7.6 ± 0.2 days while SSD and control required 10.2 ± 0.4 and 10.4 ± 0.3 days, respectively. HNE maintained a moist wound environment, were well debrided, and exhibited improved hemostatic response. Further histological examination revealed enhanced granulation tissue as compared to silver sulfadiazine and control cohorts. These results were corroborated with 3D topographical imprints of the wounds at day 14 which qualitatively showed a smoother surface. In contrast, silver sulfadiazine appeared to delay wound closure. Finally, dermal sensitization and irritation studies conducted in guinea pig and rabbits did not reveal any acute dermal side effects from HNE exposure. The cumulative data indicates nonantibiotic-based HNEs may be a promising topical treatment for the management of contaminated wounds.

Mycological study on cholesteatoma keratin obtained during primary mastoid surgery.

OBJECTIVE: Established middle-ear cleft cholesteatoma is associated with keratinous debris, which is likely to be an ideal medium for saprophytic fungal colonisation. This prospective case study aimed to explore the incidence and nature of fungal elements in cholesteatoma keratin samples obtained during primary mastoid surgery. METHODS: All cases of middle-ear cleft cholesteatoma treated with primary mastoid surgery at the El-Sahel Teaching Hospital over a seven-month period were included. Keratinous debris obtained from the mastoid antrum was subjected to mycological analysis at the Department of Medical Microbiology and Immunology, Faculty of Medicine, Cairo University. A literature search was performed to determine the clinical and pathological relevance of fungal colonisation in cholesteatoma. RESULTS: Eighteen patients underwent primary mastoid surgery for cholesteatoma (nineteen ears in total) in a seven-month period starting 30 March 2013. Patients included 13 males and 5 females, with an age range of 9 to 45 years (mean 23 years). Fungal cultures were obtained from 17 keratin samples (89 per cent). Of these, five fungal isolates belonged to the dermatophyte group (21 per cent). CONCLUSION: Fungal colonisation in middle-ear cleft cholesteatoma probably plays a significant role in disease progression. Moreover, saprophytic fungal colonisation in cholesteatoma keratin may be responsible for the fetor commonly associated with the ear discharge.

A case of direct intracranial extension of tuberculous otitis media.

We describe a very rare case of tuberculous otitis media (TOM) with direct intracranial extension. The patient was a 55-year-old man who presented to our ENT clinic for evaluation of severe headaches and right-sided otorrhea. A biopsy of granulation tissue obtained from the right external auditory canal demonstrated chronic inflammation that was suggestive of mycobacterial infection. Magnetic resonance imaging of the brain indicated intracranial extension of TOM through a destroyed tegmen mastoideum. After 2 months of antituberculous medication, the headaches and otorrhea were controlled, and the swelling in the external ear canal subsided greatly. Rarely does TOM spread intracranially. In most such cases, intracranial extension of tuberculosis occurs as the result of hematogenous or lymphogenous spread. In rare cases, direct spread through destroyed bone can occur, as it did in our patient.

High bacterial load in negative pressure wound therapy (NPWT) foams used in the treatment of chronic wounds.

No earlier study has investigated the microbiology of negative pressure wound therapy (NPWT) foam using a standardized manner. The purpose of this study is to investigate the bacterial load and microbiological dynamics in NPWT foam removed from chronic wounds (>3 months). To determine the bacterial load, a standardized size of the removed NPWT foam was sonicated. The resulting sonication fluid was cultured, and the colony-forming units (CFU) of each species were enumerated. Sixty-eight foams from 17 patients (mean age 63 years, 71% males) were investigated. In 65 (97%) foams, ≥ 1 and in 37 (54%) ≥ 2 bacterial types were found. The bacterial load remained high during NPWT treatment, ranging from 10(4) to 10(6) CFU/ml. In three patients (27%), additional type of bacteria was found in subsequent foam cultures. The mean bacterial count ± standard deviation was higher in polyvinyl alcohol foam (6.1 ± 0.5 CFU/ml) than in polyurethane (5.5 ± 0.8 CFU/ml) (p = 0.02). The mean of log of sum of CFU/ml in foam from 125 mmHg (5.5 ± 0.8) was lower than in foam from 100 mmHg pressure (5.9 ± 0.5) (p = 0.01). Concluding, bacterial load remains high in NPWT foam, and routine changing does not reduce the load.

Influence of titanium on in vitro fibroblast-Porphyromonas gingivalis interaction in peri-implantitis.

AIM: Titanium wear particles have been found in peri-implant tissues, but their role in the pathogenesis of peri-implantitis remains unclear. We aimed to determine the in vitro inflammatory responses of peri-implant granulation tissue fibroblasts (PIGFs) to titanium particles alone and in the presence of viable Porphyromonas gingivalis. MATERIALS & METHODS: Peri-implant granulation tissue fibroblasts were challenged either with TiO2 particles, P. gingivalis or a combination of TiO2 particles and P. gingivalis. Gene expression and protein production of pro-inflammatory mediators by PIGFs were measured with PCR and ELISA, respectively. RESULTS: Higher doses of TiO2 were toxic to PIGFs and in sub-toxic doses, TiO2 caused an increase in gene expression of tumour necrosis factor (TNF)-A and increased protein production of TNF-α, interleukin (IL)-6 and IL-8. A challenge with P. gingivalis alone induced gene expression of TNF-A, IL-1ß, IL-6 and IL-8. A combined challenge with TiO2 and P. gingivalis caused a stronger increase in gene expression of TNF-A and protein production of TNF-α and MCP-1 than P. gingivalis alone. CONCLUSIONS: TiO2 particles and P. gingivalis, individually, can induce pro-inflammatory responses in PIGFs. Furthermore, TiO2 particles and viable P. gingivalis further enhance gene expression and production of TNF-α by PIGFs. Therefore, Ti wear particles in the peri-implant tissues in combination with P. gingivalis infection may contribute to the pathogenesis of peri-implantitis by enhancing the inflammation in peri-implant tissues.

Gastrostomy site infections: dealing with a common problem.

The most popular method of providing appropriate nutrition for large numbers of patients with swallowing difficulties is enteral feeding. However this treatment is not without complications, one of which is localized gastrostomy site infection. This is prevented initially by decolonisation of the oropharyngeal tract with antibiotic prophylaxis prior to insertion, and systemic antibiotics post insertion. Later complications include tracking infection, which is rare but can occur. Hypergranulation of the tissue can occur around the gastrostomy tube and this can become colonised or infected leading to further problems for the patient. A good gastrostomy site care pathway plan is required to maintain a healthy site and appropriate treatment required to minimize the infection risk.

Rapid detection of bacterial DNA in mastoid granulation tissue with nested-PCR technique.

OBJECTIVE: To detect bacterial DNA in mastoid granulation tissue from patients with chronic suppurative otitis media (CSOM). MATERIAL AND METHOD: A two-step polymerase chain reaction (nested polymerase chain reaction) technique was employed. A 16s rRNA universal primer common to all bacteria was used as a bracket primer for the first step PCR reaction. Primers specific to P aeruginosa and S. aureus were then used as nested primers for the second step PCR. Products of this process were identified by DNA sequencing. RESULTS: Among 15 clinical specimens collected, five showed positive bands specific to the species P aeruginosa, and 11 showed bands specific to the genus Staphylococcus. DNA sequencing showed 99.7 to 100% accuracy for target organisms in clinical specimens with a positive signal. The average time taken to conduct the PCR procedure was about four hours CONCLUSION: The nested PCR technique described worked well, even when the size of the mastoid granulation tissue was very small.

Infected periodontal granulation tissue contains cells expressing embryonic stem cell markers. A pilot study.

The commonly practiced removal of granulation tissue during periodontal surgery, aiming to eliminate infection and optimize healing conditions, may also remove progenitor stem cells that could otherwise support periodontal regeneration. The present study aimed to investigate if cells with embryonic stem cell properties are present in periodontal granulation tissue. During the course of flap surgery inflammatory granulation tissue was obtained from four patients and five periodontal defects. Tissues were processed in a collagenase/dispase solution to release the cells. Part of the resulting suspension was processed for bacteriological analysis (IAI PadoTest 4.5), whereas the remaining cell suspension was cultured and passaged once. Upon reaching confluence, total RNA was extracted, followed by cDNA synthesis. PCR was then performed (SYBR Green-based protocols) to measure gene expression levels of Collagen type I, and embryonic stem cell markers Nanog, Oct4, Rex-1 and Sox2. Results are expressed as 2⁻Δ(Ct) values of the target gene, calibrated against a house-keeping gene (GAPDH). A high total bacterial load up to 20.6 ± 11.0×10(6) counts/mg of tissue was found. Collagen type I was strongly expressed, confirming the predominance of mesenchymal/fibroblastic cells. Among the studied embryonic stem cells markers, Nanog was most highly expressed (2.3 ± 1.2), followed by Oct4 (1.1 ± 0.5), Rex-1 (0.6 ± 0.2) and Sox2 (0.3 ± 0.2). This is the first study that demonstrates the presence of cells expressing embryonic stem cell markers among infected granulation tissue. This knowledge needs to be considered when devising future strategies to improve periodontal wound healing and regeneration.

Cytokine and matrix metalloproteinase expression in fibroblasts from peri-implantitis lesions in response to viable Porphyromonas gingivalis.

BACKGROUND AND OBJECTIVE: To assess inflammatory reactions of fibroblasts in the pathophysiology of peri-implantitis, we compared the pro-inflammatory and matrix-degrading responses of gingival and granulation tissue fibroblasts from periodontally healthy controls, peri-implantitis, and periodontitis lesions to an in vitro challenge with Porphyromonas gingivalis. METHODS: Fibroblasts from periodontally healthy, peri-implantitis and periodontitis donors were challenged with viable P. gingivalis. The inflammatory reactions of fibroblasts were analyzed before and after 6 h P. gingivalis challenge, and 2.5 and 18 h after removal of the challenge. Gene expression and induction of pro-inflammatory mediators, and matrix metalloproteinases (MMPs) were assessed by real-time polymerase chain reaction. Protein expression was measured by enzyme-linked immunosorbent assay. RESULTS: Non-challenged fibroblasts from peri-implantitis and periodontitis lesions expressed higher levels of interleukin (IL)-1ß, IL-8, and monocyte chemotactic protein (MCP)-1 than fibroblasts from periodontally healthy individuals. The P. gingivalis challenge induced expression of IL-1ß, IL-8, IL-6, MCP-1, and MMP-1 in periodontitis and peri-implantitis fibroblasts, but not in fibroblasts from periodontally healthy individuals. MMP-8 expression was higher in non-challenged peri-implantitis fibroblasts than in fibroblasts from periodontally healthy individuals. However, the P. gingivalis challenge downregulated MMP-8 gene expression in peri-implantitis fibroblasts. After removal of the P. gingivalis challenge, peri-implantitis fibroblasts sustained higher induction of IL-1ß, MCP-1, and MMP-1 compared to periodontitis fibroblasts. CONCLUSIONS: Fibroblasts from peri-implantitis and periodontitis lesions gave a more pronounced inflammatory response to the P. gingivalis challenge than fibroblasts from healthy donors. They may therefore be involved in the development of inflammation in peri-implantitis and periodontitis. Moreover, the sustained upregulation of inflammatory mediators and MMP-1 in peri-implantitis fibroblasts may play a role in the pathogenesis of peri-implantitis.

Proteolytic activity by multiple bacterial species isolated from chronic venous leg ulcers degrades matrix substrates.

BACKGROUND: A major feature of chronic wounds is the loss of tissue, with the exposure of dermal components preventing primary closure and leading to bacterial colonization. Bacterial colonization has been proposed as one of the common underlying pathologies present in chronic wounds. The objective of this exploratory study was to identify bacteria cultured from chronic venous leg ulcers and test for proteolytic activity that degrades matrix substrates. METHOD: Bacteria were isolated, cultured, and identified from six subjects (average age = 62.8 years) over 2-10 months under an approved protocol using swabs and microbiological culture media. Proteolytic activity against (a) gelatin, (b) an elastin substrate, and (c) a serine/trypsin-sensitive substrate was determined using a colorimetric plate assay with an ELISA plate reader and zymography. RESULTS: We identified 13 bacteria that expressed proteolytic activity against one or more of the tested substrates. Of these, six were Gram-positive (Staphylococcus aureus, Enterococcus faecalis, Staphylococcus epidermidis, Streptococcus agalactiae, Corynebacterium, and Streptococcus bovis) and seven were Gram-negative (Pseudomonas aeruginosa, Escherichia coli, Proteus mirabilis, Morganella morganii, Klebsiella pneumoniae, Bacteroides fragilis, and Serratia marcescens) organisms. Two of these, S. aureus and P. aeruginosa, are recognized wound pathogens. CONCLUSIONS: Multiple bacteria species isolated from colonized venous leg ulcers have the capacity to secrete proteases capable of degrading components of the extracellular matrix important for wound healing. Matrix degradation by bacteria may contribute to delays in tissue deposition and repair, suggesting that treatment of chronic wounds should include appropriate management of colonizing bacteria.

Potential implications of biofilm in chronic wounds: a case series.

Bacterial biofilm is increasingly suspected as being a significant barrier to wound healing. Bacteria predominantly attach to surfaces in their natural habitats and form biofilm; in this state they adapt to, and tolerate, the hostilities in their surrounding environment. The purpose of this clinical observational study was to consider chronic wound biofilm in relation to other factors that are implicated in wound recalcitrance, such as peripheral arterial disease, wound infection, osteomyelitis and moisture imbalance. Based on our clinical observations, it is possible that links exist between wound biofilm and other underlying pathophysiological factors, and that biofilm may also provide clues to the involvement of such factors. Recognising and managing these factors collectively may be important in addressing recalcitrance and facilitating wound progression.