Bacteria Living in Biofilms in Fluids: Could Chemical Antibiofilm Pretreatment of Culture Represent a Paradigm Shift in Diagnostics?

Biofilms are multicellular aggregates of bacteria immersed in an extracellular matrix that forms on various surfaces, including biological tissues and artificial surfaces. However, more and more reports point out the fact that even biological fluids and semifluid, such as synovial liquid, blood, urine, or mucus and feces, harbor "non-attached" biofilm aggregates of bacteria, which represent a significant phenomenon with critical clinical implications that remain to be fully investigated. In particular, biofilm aggregates in biological fluid samples have been shown to play a relevant role in bacterial count and in the overall accuracy of microbiological diagnosis. In line with these observations, the introduction in the clinical setting of fluid sample pretreatment with an antibiofilm chemical compound called dithiothreitol (DTT), which is able to dislodge microorganisms from their intercellular matrix without killing them, would effectively improve the microbiological yield and increase the sensitivity of cultural examination, compared to the current microbiological techniques. While other ongoing research continues to unveil the complexity of biofilm formation in biological fluids and its impact on infection pathogenesis and diagnosis, we here hypothesize that the routine use of a chemical antibiofilm pretreatment of fluid and semi-solid samples may lead to a paradigm shift in the microbiological approach to the diagnosis of biofilm-related infections and should be further investigated and eventually implemented in the clinical setting.

In vitro laboratory infection research in orthopaedics: Why, when, and how.

The musculoskeletal system involves multiple tissues which are constantly exposed to being exposed to various biological and mechanical stimuli. As such, isolating and studying a particular system from a complex human clinical environment is not always a realistic expectation. On top of that, recruitment limitations, in addition to the nature of orthopaedic interventions and their associated cost, sometimes preclude consideration of human trials to answer a clinical question. Therefore, in this mini review, we sought to rationalize the rapid evolution of biomedical research at a basic scientific level and explain why the perception of orthopaedic conditions has fundamentally changed over the last decades. In more detail, we highlight that the number of orthopaedic in vitro publications has soared since 1990. Last but not least, we elaborated on the minimum requirements for conducting a scientifically sound infection-related laboratory experiment to offer valuable information to clinical practitioners. We also explained the rationale behind implementing molecular biology techniques, ex vivo experiments, and artificial intelligence in this type of laboratory research.

Expert consensus on the management of patients undergoing cataract surgery: A Delphi study.

PURPOSE: To define the management of patients undergoing cataract surgery. SETTING: Panel of experts on cataract surgery and members of the Italian Association of Cataract and Refractive Surgery (AICCER) participated in the Delphi study. DESIGN: A restricted panel of experts defined the statements concerning the topic and identified a larger panel of experts who voted the statements. The statements concerned a series of practical issues concerning the management of patients undergoing cataract surgery. METHODS: An initial web round-table served to develop the statements. The larger panel was constituted by 15 experts which anonymously voted the statements, presented in a web platform, using a 5- point Likert scale. Consensus was defined as at least 80% of agreement. RESULTS: All participants completed the questionnaire. Globally, the total percentage of agreement of all statements was 90.4%. The total mean score was 4.5. Score 4 and score 5 accounted for the 27% and the 68% of the total votes, respectively. CONCLUSIONS: The participants felt they could largely agree with and approve the statements proposed by the board. In addition, the Delphi study identified some points that are highly shared and endorsed. In particular, a new model approach can be based on a seven-day course using a fixed high-potency corticosteroid combination with a broad-spectrum antibiotic. At the end of this cycle, the patient should be re-evaluated in some way to continue any treatment in the most appropriate and personalized way possible.

Strategies for Inhibition of Biofilm Formation on Silicone Rubber Voice Prostheses: A Systematic Review.

BACKGROUND: Lifetime elongation of the silicone voice rubber prostheses by inhibition of biofilm formation is a primary objective in voice restoration of laryngectomized patients. This systematic review sought to explore the existing strategies in this direction. MATERIALS: We conducted a systematic search of both in vitro and in vivo literature published in PubMed, Scopus, and Cochrane Central Register of Controlled Trials, until December 31, 2022, for published and unpublished trials assessing the strategies for inhibiting biofilm formation on silicone rubber voice prostheses, and appraised quality assessment with the modified Consolidated Standards of Reporting Trials tool. We analyzed the infection prevention capacity of the included antibacterial and antifungal agents. RESULTS: The qualitative synthesis showed that both surface modification methods and prophylactic treatment of silicone rubber voice prostheses present adequate antibiofilm activity. Of note, the majority of the suggested prosthetic surfaces were not chronically exposed to both human fluids and biofilm-forming microorganisms. CONCLUSION: Various experimental methods provide promising antibiofilm activity and, thus, possible lifespan elongation of silicone rubber voice prostheses.

Νanomaterial-Loaded Polymer Coating Prevents the In Vitro Growth of Candida albicans Biofilms on Silicone Biomaterials.

Early failure of silicone voice prostheses resulting from fungal colonization and biofilm formation poses a major concern in modern ear nose throat surgery. Therefore, developing new infection prevention techniques to prolong those implants' survivorship is crucial. We designed an in vitro laboratory study to include nanomaterial-enhanced polymer coating with a plasma spraying technique against Candida albicans growth to address this issue. The anti-biofilm effects of high- and low-dose Al2O3 nanowire and TiO2 nanoparticle coatings were studied either alone or in conjunction with each other using checkerboard testing. It was demonstrated that both nanomaterials were capable of preventing fungal biofilm formation regardless of the anti-fungal agent concentration (median absorbance for high-dose Al2O3-enhanced polymer coating was 0.176 [IQR = 0.207] versus control absorbance of 0.805 [IQR = 0.381], p = 0.003 [98% biofilm reduction]; median absorbance for high-dose TiO2-enhanced polymer coating was 0.186 [IQR = 0.024] versus control absorbance of 0.766 [IQR = 0.458], p < 0.001 [93% biofilm reduction]). Furthermore, synergy was revealed when the Bliss model was applied. According to the findings of this work, it seems that simultaneous consideration of Al2O3 and TiO2 could further increase the existing antibiofilm potential of these nanomaterials and decrease the likelihood of localized toxicity.

Is nanomaterial- and vancomycin-loaded polymer coating effective at preventing methicillin-resistant Staphylococcus aureus growth on titanium disks? An in vitro study.

PURPOSE: Periprosthetic joint infections induced by methicillin-resistant Staphylococcus aureus (MRSA) pose a major socioeconomic burden. Given the fact that MRSA carriers are at high risk for developing periprosthetic infections regardless of the administration of eradication treatment pre-operatively, the need for developing new prevention modalities is high. METHODS: The antibacterial and antibiofilm properties of vancomycin, Al2O3 nanowires, and TiO2 nanoparticles were evaluated in vitro using MIC and MBIC assays. MRSA biofilms were grown on titanium disks simulating orthopedic implants, and the infection prevention potential of vancomycin-, Al2O3 nanowire-, and TiO2 nanoparticle-supplemented Resomer® coating was evaluated against biofilm controls using the XTT reduction proliferation assay. RESULTS: Among the tested modalities, high- and low-dose vancomycin-loaded Resomer® coating yielded the most satisfactory metalwork protection against MRSA (median absorbance was 0.1705; [IQR = 0.1745] vs control absorbance 0.42 [IQR = 0.07]; p = 0.016; biofilm reduction was 100%; and 0.209 [IQR = 0.1295] vs control 0.42 [IQR = 0.07]; p < 0.001; biofilm reduction was 84%, respectively). On the other hand, polymer coating alone did not provide clinically meaningful biofilm growth prevention (median absorbance was 0.2585 [IQR = 0.1235] vs control 0.395 [IQR = 0.218]; p < 0.001; biofilm reduction was 62%). CONCLUSIONS: We advocate that apart from the well-established preventative measures for MRSA carriers, loading implants with bioresorbable Resomer® vancomycin-supplemented coating may decrease the incidence of early post-op surgical site infections with titanium implants. Of note, the payoff between localized toxicity and antibiofilm efficacy should be considered when loading polymers with highly concentrated antimicrobial agents.

Dithiotreitol pre-treatment of synovial fluid samples improves microbiological counts in peri-prosthetic joint infection.

PURPOSE: Synovial fluid cultures of periprosthetic joint infections (PJI) may be limited by bacteria living in the fluids as biofilm-aggregates. The antibiofilm pre-treatment of synovial fluids with dithiotreitol (DTT) could improve bacterial counts and microbiological early stage diagnosis in patients with suspected PJI. METHODS: Synovial fluids collected from 57 subjects, affected by painful total hip or knee replacement, were divided into two aliquots, one pre-treated with DTT and one with normal saline. All samples were plated for microbial counts. Sensitivity of cultural examination and bacterial counts of pre-treated and control samples were then calculated and statistically compared. RESULTS: Dithiothreitol pre-treatment led to a higher number of positive samples, compared to controls (27 vs 19), leading to a statistically significant increase in the sensitivity of the microbiological count examination from 54.3 to 77.1% and in colony-forming units count from 1884 ± 2.129 CFU/mL with saline pre-treatment to 20.442 ± 19.270 with DTT pre-treatment (P = 0.02). CONCLUSIONS: To our knowledge, this is the first report showing the ability of a chemical antibiofilm pre-treatment to increase the sensitivity of microbiological examination in the synovial fluid of patients with peri-prosthetic joint infection. If confirmed by larger studies, this finding may have a significant impact on routine microbiological procedures applied to synovial fluids and brings further support to the key role of bacteria living in biofilm-formed aggregates in joint infections.

Viral infections in orthopedics: A systematic review and classification proposal.

BACKGROUND: Although the impact of microbial infections on orthopedic clinical outcomes is well recognized, the influence of viral infections on the musculoskeletal system might have been underestimated. AIM: To systematically review the available evidence on risk factors and musculoskeletal manifestations following viral infections and to propose a pertinent classification scheme. METHODS: We searched MEDLINE, Cochrane Central Register of Controlled Trials (CENTRAL), the Reference Citation Analysis (RCA), and Scopus for completed studies published before January 30, 2021, to evaluate risk factors and bone and joint manifestations of viral infection in animal models and patient registries. Quality assessment was performed using SYRCLE's risk of bias tool for animal studies, Moga score for case series, Wylde score for registry studies, and Newcastle-Ottawa Scale for case-control studies. RESULTS: Six human and four animal studies were eligible for inclusion in the qualitative synthesis. Hepatitis C virus was implicated in several peri- and post-operative complications in patients without cirrhosis after major orthopedic surgery. Herpes virus may affect the integrity of lumbar discs, whereas Ross River and Chikungunya viruses provoke viral arthritis and bone loss. CONCLUSION: Evidence of moderate strength suggested that viruses can cause moderate to severe arthritis and osteitis. Risk factors such as pre-existing rheumatologic disease contributed to higher disease severity and duration of symptoms. Therefore, based on our literature search, the proposed clinical and pathogenetic classification scheme is as follows: (1) Viral infections of bone or joint; (2) Active bone and joint inflammatory diseases secondary to viral infections in other organs or tissues; and (3) Viral infection as a risk factor for post-surgical bacterial infection.

Randomized Double-Blind Crossover Study for Evaluating a Probiotic Mixture on Gastrointestinal and Behavioral Symptoms of Autistic Children.

Autism spectrum disorders (ASDs) represent a diagnostic challenge with a still partially uncertain etiology, in which genetic and environmental factors have now been assessed. Among the hypotheses underlying the involvement of biological and environmental factors, the gut-brain axis is of particular interest in autism spectrum disorders. Several studies have highlighted the related incidence of particular gastrointestinal symptoms (GISs) in children suffering from ASDs. Probiotics have shown success in treating several gastrointestinal dysbiotic disorders; therefore, it is plausible to investigate whether they can alleviate behavioral symptoms as well. On these bases, a randomized double-blind crossover study with a placebo was conducted, evaluating the effects of a mixture of probiotics in a group of 61 subjects aged between 24 months and 16 years old with a diagnosis of ASD. Behavioral evaluation was performed through the administration of a questionnaire including a Parenting Stress Index (PSI) test and the Vineland Adaptive Behavior Scale (VABS). The Psycho-Educational Profile and the Autism Spectrum Rating Scale (ASRS) were also evaluated. Microbial composition analyses of fecal samples of the two groups was also performed. The study showed significant improvements in GISs, communication skills, maladaptive behaviors, and perceived parental stress level after the administration of probiotics. Microbiome alpha diversity was comparable between treatment arms and no significant differences were found, although beta diversity results were significantly different in the treatment group between T0 and T1 time points. Streptococcus thermophilus, Bifidobacterium longum, Limosilactobacillus fermentum, and Ligilactobacillus salivarius species were identified as some of the most discriminant taxa positively associated with T1 samples. This preliminary study corroborates the relationship between intestinal microbiota and ASD recently described in the literature.

Infectious disease specialists and teamwork strategies worldwide: the World Association against Infection in Orthopedics and Trauma (WAIOT) and SICOT continue to cooperate in fighting musculoskeletal infections.

Bone and joint infections are associated with a devastating global burden. The successful treatment of these infections requires a multidisciplinary approach between orthopedic surgeons and experts of different disciplines. This multidisciplinary approach has gained ground over the past decades in modern infection units as a more effective treatment strategy, yielding better outcomes regarding infection eradication rates, length of hospital stay, and overall cost of treatments. Additionally, preventing and managing musculoskeletal infections requires strong connections between medical associations, biological laboratories, and the pharmaceutical industry worldwide. In this context, SICOT and World Association against Infection in Orthopaedics and Trauma (WAIOT) relationships have been increasing. The present editorial article discusses the multidisciplinary approach for managing bone and joint infections worldwide, explores the controversies in practices in terms of training, area of expertise, and extent of clinical involvement, and emphasizes the role of societies in research, prevention and management of musculoskeletal infections. The purpose is to acknowledge what orthopedics can obtain from specialists dealing with bone and joint infections and to consolidate their practice to provide the best care for orthopedic patients.

A post hoc analysis on the effects of a probiotic mixture on asthma exacerbation frequency in schoolchildren.

The present randomised, placebo-controlled, double-blind study showed that a probiotic mixture significantly reduced the number of asthma exacerbations in schoolchildren https://bit.ly/382LYKV.

Metal Hypersensitivity or Missed Periprosthetic Joint Infection? A Critical Review.

The World Association Against Infection in Orthopedics And Trauma (W.A.I.O.T.) Study Group on Bone And Joint Infection Definitions Metal hypersensitivity (MHS) has been investigated by several authors as a possible reason for painful total joint arthroplasty, with controversial results. Periprosthetic joint infection (PJI) is another possible source of unexplained pain and implant failure that may be difficult to diagnose if not properly investigated. We performed this critical review to assess whether the current literature on MHS includes an adequate diagnostic workup to discern metal allergy from PJI. The results of this review highlight the importance of assessing patients for PJI before making a diagnosis of MHS and emphasize that the methods currently used to exclude PJI are substantially inadequate. Therefore, well-designed clinical trials with adequate diagnostic protocols and definitions of PJI that can differentiate MHS from low-grade PJI are needed. [Orthopedics. 2022;45(2):e73-e78.].

The Probiotics in Pediatric Asthma Management (PROPAM) Study in the Primary Care Setting: A Randomized, Controlled, Double-Blind Trial with Ligilactobacillus salivarius LS01 (DSM 22775) and Bifidobacterium breve B632 (DSM 24706).

BACKGROUND: Type-2 inflammation commonly marks asthma in childhood. Also, gut and lung dysbiosis is detectable in patients with asthma. Strain-related probiotic supplementation may restore a physiological immune response, dampen airway inflammation, and repair dysbiosis. Therefore, the probiotics in pediatric asthma management (PROPAM) study is aimed at demonstrating that Ligilactobacillus salivarius LS01 (DSM 22775) and Bifidobacterium breve B632 (DSM 24706) mixture could reduce asthma exacerbations in children, followed in a primary care setting. METHODS: The study was randomized, placebo-controlled, and double-blind. It involved 11 Italian primary care pediatricians. The probiotic mixture (containing Ligilactobacillus salivarius LS01 1 × 109 live cells and Bifidobacterium breve B632 1 × 109 live cells) or placebo was taken twice daily (1 sachet in the morning and 1 in the evening) for eight weeks and subsequently once daily for a further eight weeks. Outcomes included number, severity, and duration of asthma exacerbations, intensity of maintenance and as need treatments, and safety. RESULTS: The per-protocol population included 422 children (mean age seven years, 240 males and 182 females). The probiotic mixture significantly reduced the number of asthmatic exacerbations (OR = 3.17). In addition, the number of children with two exacerbations was less than a third in the active group (OR = 3.65). CONCLUSIONS: This PROPAM study demonstrated that probiotic strains Ligilactobacillus salivarius LS01 (DSM 22775) and Bifidobacterium breve B632 (DSM 24706) were safe and significantly reduced by more than a third the frequency of asthma exacerbations. At present, the first-line treatment of asthma is still drug-based, but specific strains of probiotics may be auxiliary remedies.

In vitro Inhibition of Biofilm Formation on Silicon Rubber Voice Prosthesis: Α Systematic Review and Meta-Analysis.

INTRODUCTION: Biofilm formation on voice prostheses is the primary reason for their premature implant dysfunction. Multiple strategies have been proposed over the last decades to achieve inhibition of biofilm formation on these devices. The purpose of this study was to assess the results of the available in vitro biofilm inhibition modalities on silicone rubber voice prostheses. METHODS: We conducted a systematic search in PubMed, Embase, and the Cochrane Central Register of Controlled Trials databases up to February 29, 2020. A total of 33 in vitro laboratory studies investigating the efficacy of different coating methods against Candida, Staphylococcus, Streptococcus, Lactobacilli, and Rothia biofilm growth on silicone rubber medical devices were included. Subgroup analysis linked to the type of prevention modality was carried out, and quality assessment was performed with the use of the modified CONSORT tool. RESULTS: Data from 33 studies were included in qualitative analysis, of which 12 qualified for quantitative analysis. For yeast biofilm formation assessment, there was a statistically significant difference in favor of the intervention group (standardized mean difference [SMD] = -1.20; 95% confidence interval [CI] [-1.73, -0.66]; p < 0.0001). Subgroup analysis showed that combined methods (active and passive surface modification) are the most effective for biofilm inhibition in yeast (SMD = -2.53; 95% CI [-4.02, -1.03]; p = 0.00001). No statistically significant differences between intervention and control groups were shown for bacterial biofilm inhibition (SMD = -0.09; 95% CI [-0.68, 0.46]; p = 0.65), and the results from the subgroup analysis found no notable differences between the surface modification methods. After analyzing data on polymicrobial biofilms, a statistically significant difference in favor of prevention methods in comparison with the control group was detected (SMD = -2.59; 95% CI [-7.48, 2.31]; p = 0.30). CONCLUSIONS: The meta-analysis on biofilm inhibition demonstrated significant differences in favor of yeast biofilm inhibition compared to bacteria. A stronger inhibition with the application of passive or combined active and passive surface modification techniques was reported.

A Journey on the Skin Microbiome: Pitfalls and Opportunities.

The human skin microbiota is essential for maintaining homeostasis and ensuring barrier functions. Over the years, the characterization of its composition and taxonomic diversity has reached outstanding goals, with more than 10 million bacterial genes collected and cataloged. Nevertheless, the study of the skin microbiota presents specific challenges that need to be addressed in study design. Benchmarking procedures and reproducible and robust analysis workflows for increasing comparability among studies are required. For various reasons and because of specific technical problems, these issues have been investigated in gut microbiota studies, but they have been largely overlooked for skin microbiota. After a short description of the skin microbiota, the review tackles methodological aspects and their pitfalls, covering NGS approaches and high throughput culture-based techniques. Recent insights into the "core" and "transient" types of skin microbiota and how the manipulation of these communities can prevent or combat skin diseases are also covered. Finally, this review includes an overview of the main dermatological diseases, the changes in the microbiota composition associated with them, and the recommended skin sampling procedures. The last section focuses on topical and oral probiotics to improve and maintain skin health, considering their possible applications for skin diseases.

Deep Transcranial Magnetic Stimulation Affects Gut Microbiota Composition in Obesity: Results of Randomized Clinical Trial.

Growing evidence highlights the crucial role of gut microbiota in affecting different aspects of obesity. Considering the ability of deep transcranial magnetic stimulation (dTMS) to modulate the cortical excitability, the reward system, and, indirectly, the autonomic nervous system (ANS), we hypothesized a potential role of dTMS in affecting the brain-gut communication pathways, and the gut microbiota composition in obesity. In a hospital setting, 22 subjects with obesity (5 M, 17 F; 44.9 ± 2.2 years; BMI 37.5 ± 1.0 kg/m2) were randomized into three groups receiving 15 sessions (3 per week for 5 weeks) of high frequency (HF), low frequency (LF) dTMS, or sham stimulation. Fecal samples were collected at baseline and after 5 weeks of treatment. Total bacterial DNA was extracted from fecal samples using the QIAamp DNA Stool Mini Kit (Qiagen, Italy) and analyzed by a metagenomics approach (Ion Torrent Personal Genome Machine). After 5 weeks, a significant weight loss was found in HF (HF: -4.1 ± 0.8%, LF: -1.9 ± 0.8%, sham: -1.3 ± 0.6%, p = 0.042) compared to LF and sham groups, associated with a decrease in norepinephrine compared to baseline (HF: -61.5 ± 15.2%, p < 0.01; LF: -31.8 ± 17.1%, p < 0.05; sham: -35.8 ± 21.0%, p > 0.05). Furthermore, an increase in Faecalibacterium (+154.3% vs. baseline, p < 0.05) and Alistipes (+153.4% vs. baseline, p < 0.05) genera, and a significant decrease in Lactobacillus (-77.1% vs. baseline, p < 0.05) were found in HF. Faecalibacterium variations were not significant compared to baseline in the other two groups (LF: +106.6%, sham: +27.6%; p > 0.05) as well as Alistipes (LF: -54.9%, sham: -15.1%; p > 0.05) and Lactobacillus (LF: -26.0%, sham: +228.3%; p > 0.05) variations. Norepinephrine change significantly correlated with Bacteroides (r2 = 0.734; p < 0.05), Eubacterium (r2 = 0.734; p < 0.05), and Parasutterella (r2 = 0.618; p < 0.05) abundance variations in HF. In conclusion, HF dTMS treatment revealed to be effective in modulating gut microbiota composition in subjects with obesity, reversing obesity-associated microbiota variations, and promoting bacterial species representative of healthy subjects with anti-inflammatory properties.