Reduced Glycolysis and Cytotoxicity in Staphylococcus aureus Isolates from Chronic Rhinosinusitis as Strategies for Host Adaptation.

Chronic rhinosinusitis (CRS) is a multifactorial infection of the nasal cavity and sinuses. In this study, nasal swabs from control donors (N = 128) and patients with CRS (N = 246) were analysed. Culture methods and metagenomics revealed no obvious differences in the composition of the bacterial communities between the two groups. However, at the functional level, several metabolic pathways were significantly enriched in the CRS group compared to the control group. Pathways such as carbohydrate transport metabolism, ATP synthesis, cofactors and vitamins, photosynthesis and transcription were highly enriched in CRS. In contrast, pathways related to lipid metabolism were more representative in the control microbiome. As S. aureus is one of the main species found in the nasal cavity, staphylococcal isolates from control and CRS samples were analysed by microarray and functional assays. Although no significant genetic differences were detected by microarray, S. aureus from CRS induced less cytotoxicity to lung cells and lower rates of glycolysis in host cells than control isolates. These results suggest the differential modulation of staphylococcal virulence by the environment created by other microorganisms and their interactions with host cells in control and CRS samples. These changes were reflected in the differential expression of cytokines and in the expression of Agr, the most important quorum-sensing regulator of virulence in S. aureus. In addition, the CRS isolates remained stable in their cytotoxicity, whereas the cytotoxic activity of S. aureus isolated from control subjects decreased over time during in vitro passage. These results suggest that host factors influence the virulence of S. aureus and promote its adaptation to the nasal environment during CRS.

Thoracic adipose tissue contributes to severe virus infection of the lung.

OBJECTIVE: Obesity is an independent risk factor for severe influenza virus and COVID-19 infections. There might be an interplay between adipose tissue and respiratory pathogens, although the mechanism is unknown. Proinflammatory factors secreted by the adipose tissue are often discussed to serve as indirect contributor to virus infection. However, the direct potential of adipose tissue to serve as a viral niche has not yet been investigated. METHODS: Two murine obesity models (DIO and ob/ob) were infected with influenza A virus (IAV) and monitored for 3 weeks. p.i. Lung and adipose tissue were harvested, and the viral load was analysed. Direct replication of IAV in vitro was investigated in human derived primary adipocytes and macrophages. The indirect impact of the secretory products of adipocytes during infection was analysed in a co-culture system with lung fibroblasts. Moreover, lung and adipose tissue was harvested from deceased patients infected with SARS-CoV-2 omicron variant. Additionally, replication of SARS-CoV-2 alpha, delta, and omicron variants was investigated in vitro in adipocytes and macrophages. RESULTS: Both murine obesity models presented high IAV titers compared to non-obese mice. Interestingly, adipose tissue adjacent to the lungs was a focal point for influenza virus replication in mice. We further detected IAV replication and antiviral response in human adipocytes. Co-cultivation of adipocytes and lung fibroblasts led to increased IL-8 concentration during infection. Though we observed SARS-CoV-2 in the thoracic adipose tissue of COVID-19 patients, no active replication was found in adipocytes in vitro. However, SARS-CoV-2 was detected in the macrophages and this finding was associated with increased inflammation. CONCLUSIONS: Our study revealed that thoracic adipose tissue contributes to respiratory virus infection. Besides indirect induction of proinflammatory factors during infection, adipocytes and macrophages within the tissue can directly support viral replication.

Development of a rapid diagnostic test based on loop-mediated isothermal amplification to identify the most frequent non-typhoidal Salmonella serovars from culture.

Identification of Salmonella serovars is performed by conventional seroagglutination or sequencing. These methods are labor-intensive and require technical experience. An easy-to-perform assay allowing the timely identification of the most common non-typhoidal serovars (NTS) is needed. In this study, a molecular assay based on loop-mediated isothermal amplification (LAMP) targeting specific gene sequences of Salmonella Enteritidis, S. Typhimurium, S. Infantis, S. Derby, and S. Choleraesuis has been developed for rapid serovar identification from cultured colonies. A total of 318 Salmonella strains and 25 isolates of other Enterobacterales species that served as negative controls were analyzed. All S. Enteritidis (n = 40), S. Infantis (n = 27), and S. Choleraesuis (n = 11) strains were correctly identified. Seven out of 104 S. Typhimurium and 10 out of 38 S. Derby strains missed a positive signal. Cross-reactions of the gene targets were only rarely observed and restricted to the S. Typhimurium primer set (5 false-positives). Sensitivity and specificity of the assay compared to seroagglutination were as follows: 100% and 100% for S. Enteritidis, 93.3% and 97.7% for S. Typhimurium, 100% and 100% for S. Infantis, 73.7% and 100% for S. Derby, and 100% and 100% for S. Choleraesuis, respectively. With results available in just a few minutes of hands-on time and a test run time of 20 min, the LAMP assay developed here may be a useful tool for the rapid identification of common Salmonella NTS in daily routine diagnostics.

Non-patient-related SARS-CoV-2 exposure from colleagues and household members poses the highest infection risk for hospital employees in a German university hospital: follow-up of the prospective Co-HCW seroprevalence study.

PURPOSE: The Co-HCW study is a prospective, longitudinal, single-center observational study that aims to assess the SARS-CoV-2 seroprevalence and infection status in staff members of Jena University Hospital (JUH) in Jena, Germany. METHODS: This follow-up study covers the observation period from 19th May 2020 to 22nd June 2021. At each of the three voluntary study visits, participants filled out a questionnaire regarding their SARS-CoV-2 exposure and provided serum samples to detect specific SARS-CoV-2 antibodies. Participants who were tested positive for antibodies against nucleocapsid and/or spike protein without previous vaccination and/or reported a positive SARS-CoV-2 PCR test were regarded to have been infected with SARS-CoV-2. Multivariable logistic regression modeling was applied to identify potential risk factors for infected compared to non-infected participants. RESULTS: Out of 660 participants that were included during the first study visit, 406 participants (61.5%) were eligible for the final analysis as their COVID-19 risk area (high-risk n = 76; intermediate-risk n = 198; low-risk n = 132) did not change during the study. Forty-four participants [10.8%, 95% confidence interval (95%CI) 8.0-14.3%] had evidence of a current or past SARS-CoV-2 infection detected by serology (n = 40) and/or PCR (n = 28). No association between SARS-CoV-2 infection and the COVID-19 risk group according to working place was detected. However, exposure to a SARS-CoV-2 positive household member [adjusted OR (AOR) 4.46, 95% CI 2.06-9.65] or colleague (AOR 2.30, 95%CI 1.10-4.79) was found to significantly increase the risk of a SARS-CoV-2 infection. CONCLUSION: Our results demonstrate that non-patient-related SARS-CoV-2 exposure posed the highest infection risk for hospital staff members of JUH.

The Local Anaesthetic Procaine Prodrugs ProcCluster® and Procaine Hydrochloride Impair SARS-CoV-2 Replication and Egress In Vitro.

As vaccination efforts against SARS-CoV-2 progress in many countries, there is still an urgent need for efficient antiviral treatment strategies for those with severer disease courses, and lately, considerable efforts have been undertaken to repurpose existing drugs as antivirals. The local anaesthetic procaine has been investigated for antiviral properties against several viruses over the past decades. Here, we present data on the inhibitory effect of the procaine prodrugs ProcCluster® and procaine hydrochloride on SARS-CoV-2 infection in vitro. Both procaine prodrugs limit SARS-CoV-2 progeny virus titres as well as reduce interferon and cytokine responses in a proportional manner to the virus load. The addition of procaine during the early stages of the SARS-CoV-2 replication cycle in a cell culture first limits the production of subgenomic RNA transcripts, and later affects the replication of the viral genomic RNA. Interestingly, procaine additionally exerts a prominent effect on SARS-CoV-2 progeny virus release when added late during the replication cycle, when viral RNA production and protein production are already largely completed.

The Controversial Effect of Antibiotics on Methicillin-Sensitive S. aureus: A Comparative In Vitro Study.

Methicillin-sensitive Staphylococcus (S.) aureus (MSSA) bacteremia remains a global challenge, despite the availability of antibiotics. Primary treatments include ß-lactam agents such as cefazolin and flucloxacillin. Ongoing discussions have focused on the potential synergistic effects of combining these agents with rifampicin or fosfomycin to combat infections associated with biofilm formation. Managing staphylococcal infections is challenging due to antibacterial resistance, biofilms, and S. aureus's ability to invade and replicate within host cells. Intracellular invasion shields the bacteria from antibacterial agents and the immune system, often leading to incomplete bacterial clearance and chronic infections. Additionally, S. aureus can assume a dormant phenotype, known as the small colony variant (SCV), further complicating eradication and promoting persistence. This study investigated the impact of antibiotic combinations on the persistence of S. aureus 6850 and its stable small colony variant (SCV strain JB1) focusing on intracellular survival and biofilm formation. The results from the wild-type strain 6850 demonstrate that ß-lactams combined with RIF effectively eliminated biofilms and intracellular bacteria but tend to select for SCVs in planktonic culture and host cells. Higher antibiotic concentrations were associated with an increase in the zeta potential of S. aureus, suggesting reduced membrane permeability to antimicrobials. When using the stable SCV mutant strain JB1, antibiotic combinations with rifampicin successfully cleared planktonic bacteria and biofilms but failed to eradicate intracellular bacteria. Given these findings, it is reasonable to report that ß-lactams combined with rifampicin represent the optimal treatment for MSSA bacteremia. However, caution is warranted when employing this treatment over an extended period, as it may elevate the risk of selecting for small colony variants (SCVs) and, consequently, promoting bacterial persistence.

Insights into S. aureus-Induced Bone Deformation in a Mouse Model of Chronic Osteomyelitis Using Fluorescence and Raman Imaging.

Osteomyelitis is an infection of the bone that is often difficult to treat and causes a significant healthcare burden. Staphylococcus aureus is the most common pathogen causing osteomyelitis. Osteomyelitis mouse models have been established to gain further insights into the pathogenesis and host response. Here, we use an established S. aureus hematogenous osteomyelitis mouse model to investigate morphological tissue changes and bacterial localization in chronic osteomyelitis with a focus on the pelvis. X-ray imaging was performed to follow the disease progression. Six weeks post infection, when osteomyelitis had manifested itself with a macroscopically visible bone deformation in the pelvis, we used two orthogonal methods, namely fluorescence imaging and label-free Raman spectroscopy, to characterise tissue changes on a microscopic scale and to localise bacteria in different tissue regions. Hematoxylin and eosin as well as Gram staining were performed as a reference method. We could detect all signs of a chronically florid tissue infection with osseous and soft tissue changes as well as with different inflammatory infiltrate patterns. Large lesions dominated in the investigated tissue samples. Bacteria were found to form abscesses and were distributed in high numbers in the lesion, where they could occasionally also be detected intracellularly. In addition, bacteria were found in lower numbers in surrounding muscle tissue and even in lower numbers in trabecular bone tissue. The Raman spectroscopic imaging revealed a metabolic state of the bacteria with reduced activity in agreement with small cell variants found in other studies. In conclusion, we present novel optical methods to characterise bone infections, including inflammatory host tissue reactions and bacterial adaptation.

Salivary Pellicle Formed on Dental Composites Evaluated by Mass Spectrometry-An In Situ Study.

(1) Background: In the oral environment, sound enamel and dental restorative materials are immediately covered by a pellicle layer, which enables bacteria to attach. For the development of new materials with repellent surface functions, information on the formation and maturation of salivary pellicles is crucial. Therefore, the present in situ study aimed to investigate the proteomic profile of salivary pellicles formed on different dental composites. (2) Methods: Light-cured composite and bovine enamel samples (controls) were exposed to the oral cavity for 30, 90, and 120 min. All samples were subjected to optical and mechanical profilometry, as well as SEM surface evaluation. Acquired pellicles and unstimulated whole saliva samples were analyzed by SELDI-TOF-MS. The significance was determined by the generalized estimation equation and the post-hoc bonferroni adjustment. (3) Results: SEM revealed the formation of homogeneous pellicles on all test and control surfaces. Profilometry showed that composite surfaces tend to be of higher roughness compared to enamel. SELDI-TOF-MS detected up to 102 different proteins in the saliva samples and up to 46 proteins in the pellicle. Significant differences among 14 pellicle proteins were found between the composite materials and the controls. (4) Conclusions: Pellicle formation was material- and time-dependent. Proteins differed among the composites and to the control.

Staphylococcus aureus controls eicosanoid and specialized pro-resolving mediator production via lipoteichoic acid.

Staphylococcus aureus causes severe infections associated with inflammation, such as sepsis or osteomyelitis. Inflammatory processes are regulated by distinct lipid mediators (LMs) but how their biosynthetic pathways are orchestrated in S. aureus infections is elusive. We show that S. aureus strikingly not only modulates pro-inflammatory, but also inflammation-resolving LM pathways in murine osteomyelitis and osteoclasts as well as in human monocyte-derived macrophages (MDMs) with different phenotype. Targeted LM metabololipidomics using ultra-performance liquid chromatography-tandem mass spectrometry revealed massive generation of LM with distinct LM signature profiles in acute and chronic phases of S. aureus-induced murine osteomyelitis in vivo. In human MDM, S. aureus elevated cyclooxygenase-2 (COX-2) and microsomal prostaglandin E2  synthase-1 (mPGES-1), but impaired the levels of 15-lipoxygenase-1 (15-LOX-1), with respective changes in LM signature profiles initiated by these enzymes, that is, elevated PGE2 and impaired specialized pro-resolving mediators, along with reduced M2-like phenotypic macrophage markers. The cell wall component, lipoteichoic acid (LTA), mimicked the impact of S. aureus elevating COX-2/mPGES-1 expression via NF-κB and p38 MAPK signalling in MDM, while the impairment of 15-LOX-1 correlates with reduced expression of Lamtor1. In conclusion, S. aureus dictates LM pathways via LTA resulting in a shift from anti-inflammatory M2-like towards pro-inflammatory M1-like LM signature profiles.

SARS-CoV-2 causes severe epithelial inflammation and barrier dysfunction.

Infections with SARS-CoV-2 can be asymptomatic, but they can also be accompanied by a variety of symptoms that result in mild to severe coronavirus disease-19 (COVID-19) and are sometimes associated with systemic symptoms. Although the viral infection originates in the respiratory system, it is unclear how the virus can overcome the alveolar barrier, which is observed in severe COVID-19 disease courses. To elucidate the viral effects on the barrier integrity and immune reactions, we used mono-cell culture systems and a complex human chip model composed of epithelial, endothelial, and mononuclear cells. Our data show that SARS-CoV-2 efficiently infected epithelial cells with high viral loads and inflammatory response, including interferon expression. By contrast, the adjacent endothelial layer was neither infected nor did it show productive virus replication or interferon release. With prolonged infection, both cell types were damaged, and the barrier function was deteriorated, allowing the viral particles to overbear. In our study, we demonstrate that although SARS-CoV-2 is dependent on the epithelium for efficient replication, the neighboring endothelial cells are affected, e.g., by the epithelial cytokines or components induced during infection, which further results in the damage of the epithelial/endothelial barrier function and viral dissemination.IMPORTANCESARS-CoV-2 challenges healthcare systems and societies worldwide in unprecedented ways. Although numerous new studies have been conducted, research to better understand the molecular pathogen-host interactions are urgently needed. For this, experimental models have to be developed and adapted. In the present study we used mono cell-culture systems and we established a complex chip model, where epithelial and endothelial cells are cultured in close proximity. We demonstrate that epithelial cells can be infected with SARS-CoV-2, while the endothelium did not show any infection signs. Since SARS-CoV-2 is able to establish viremia, the link to thromboembolic events in severe COVID-19 courses is evident. However, whether the endothelial layer is damaged by the viral pathogens or whether other endothelial-independent homeostatic factors are induced by the virus is essential for understanding the disease development. Therefore, our study is important as it demonstrates that the endothelial layer could not be infected by SARS-CoV-2 in our in vitro experiments, but we were able to show the destruction of the epithelial-endothelial barrier in our chip model. From our experiments we can assume that virus-induced host factors disturbed the epithelial-endothelial barrier function and thereby promote viral spread.

Neutralization of the Staphylococcus aureus Panton-Valentine leukocidin by African and Caucasian sera.

BACKGROUND: The prevalence of Staphylococcus aureus isolates carrying the Panton-Valentine leukocidin (PVL) gene is higher in Africa (≈50%) compared to Europe (< 5%). The study aimed to measure anti-PVL-antibodies in Africans and Germans in a multi-center study and to test whether detected antibodies can neutralize the cytotoxic effect of PVL on polymorphonuclear leukocytes (PMNs). METHODS: Sera from asymptomatic Africans (n = 22, Nigeria, Gabon) and Caucasians (n = 22, Germany) were used to quantify antibody titers against PVL and α-hemolysin (in arbitrary units [AU]) by ELISA. PMNs from one African and German donor were exposed to 5 nM recombinant PVL to measure the neutralizing effect of serial dilutions of pooled sera from African and Caucasian participants, or donor sera at 0.625 and 2.5% (v/v). RESULTS: Anti-PVL-antibodies were significantly higher in Africans than in Germans (1.9 vs. 0.7 AU, p < 0.0001). The pooled sera from the study participants neutralized the cytotoxic effect of PVL on African and German PMNs in a dose dependent manner. Also, neutralization of PVL on PMNs from the African and German donors had a stronger effect with African sera (half-maximal inhibitory concentration (IC50) = 0.27 and 0.47%, respectively) compared to Caucasian sera (IC50 = 3.51 and 3.59% respectively). CONCLUSION: Africans have higher levels of neutralizing anti-PVL-antibodies. It remains unclear if or at what level these antibodies protect against PVL-related diseases.

Performance of the eazyplex® BloodScreen GN as a simple and rapid molecular test for identification of Gram-negative bacteria from positive blood cultures.

The LAMP-based eazyplex® BloodScreen GN was evaluated for the detection of frequent Gram-negatives directly from positive blood culture (BC) bottles. A total of 449 BCs were analyzed. Sensitivities and specificities were 100% and 100% for Escherichia coli, 95.7% and 100% for Klebsiella pneumoniae, 100% and 100% for blaCTX-M, 100% and 100% for Klebsiella oxytoca, 100% and 99% for Proteus mirabilis, and 100% and 99.8% for Pseudomonas aeruginosa, respectively. The time to result ranged from 8 to 16 min, plus about 6 min for sample preparation. The eazyplex® BloodScreen GN is a reliable molecular assay for rapid BC testing.

Complement involvement in bone homeostasis and bone disorders.

An integral part of innate immunity is the complement system, a defence system, consisting of fluid-phase and cell surface-bound proteins. Its role to ensure adequate responses to danger factors and thus promoting host defence against pathogens has been well described already for decades. Recently, numerous further reaching functions of complement have been discovered, among these are tissue homeostasis and regeneration, also with respect to the skeletal system. The influence of complement activation on bone was recognised first in pathological conditions of inflamed bone tissue and surrounding areas, observed, for example, in rheumatoid arthritis and osteoarthritis. Greatly enhanced levels of complement proteins were detected in synovial fluids and sera of arthritic patients compared to healthy individuals. Additionally, complement-mediated signalling was shown to modulate periodontitis disease development and progression. Periodontitis is an infectious condition of the periodontium, which involves severe bone loss. Moreover, the complement system critically modulates bone regeneration and healing outcome after fracture. This is seen in uneventful fracture healing, but particularly under severe inflammatory conditions induced by an additional traumatic injury. Therefore, complement activation plays an important role in both sterile and non-sterile inflammatory conditions of the bone, which will be addressed here in respect of findings in bone fractures, arthritides, periodontitis and osteomyelitis. Importantly, complement proteins are thought to be critical not simply in the states of an activated immune system, but also for bone growth during physiological development and bone homeostasis, given for example their presence in long-bone growth-plate cartilage. Furthermore, bone-cell development from precursor cells and bone-cell metabolism and communication, for example, between bone-forming osteoblasts and bone-resorbing osteoclasts, are dependent on or even critically influenced by the presence of complement proteins and complement-mediated signalling. The present review summarises the current view on the role of the complement cascade on bone, both under homeostatic physiological conditions and under inflammatory and infectious conditions, which strongly affect the bone and skeletal health. Furthermore, this review addresses the potential and the feasibility of therapeutic interventions involving the complement cascade, derived from experimental and clinical data. Modulating the complement system could help in the future to reduce bone infections, ensure a balanced bone turnover and to generally improve skeletal health.

D,L-Lysine-Acetylsalicylate + Glycine (LASAG) Reduces SARS-CoV-2 Replication and Shows an Additive Effect with Remdesivir.

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) causing the coronavirus disease-19 (COVID-19) is still challenging healthcare systems and societies worldwide. While vaccines are available, therapeutic strategies are developing and need to be adapted to each patient. Many clinical approaches focus on the repurposing of approved therapeutics against other diseases. However, the efficacy of these compounds on viral infection or even harmful secondary effects in the context of SARS-CoV-2 infection are sparsely investigated. Similarly, adverse effects of commonly used therapeutics against lifestyle diseases have not been studied in detail. Using mono cell culture systems and a more complex chip model, we investigated the effects of the acetylsalicylic acid (ASA) salt D,L-lysine-acetylsalicylate + glycine (LASAG) on SARS-CoV-2 infection in vitro. ASA is commonly known as Aspirin® and is one of the most frequently used medications worldwide. Our data indicate an inhibitory effect of LASAG on SARS-CoV-2 replication and SARS-CoV-2-induced expression of pro-inflammatory cytokines and coagulation factors. Remarkably, our data point to an additive effect of the combination of LASAG and the antiviral acting drug remdesivir on SARS-CoV-2 replication in vitro.

Ruling out COVID-19 by chest CT at emergency admission when prevalence is low: the prospective, observational SCOUT study.

BACKGROUND: It is essential to avoid admission of patients with undetected corona virus disease 2019 (COVID-19) to hospitals' general wards. Even repeated negative reverse transcription polymerase chain reaction (RT-PCR) results do not rule-out COVID-19 with certainty. The study aimed to evaluate a rule-out strategy for COVID-19 using chest computed tomography (CT) in adults being admitted to the emergency department and suspected of COVID-19. METHODS: In this prospective, single centre, diagnostic accuracy cohort study, consecutive adults (≥ 18 years) presenting with symptoms consistent with COVID-19 or previous contact to infected individuals, admitted to the emergency department and supposed to be referred to general ward were included in March and April 2020. All participants underwent low-dose chest CT. RT-PCR- and specific antibody tests were used as reference standard. Main outcome measures were sensitivity and specificity of chest CT. Predictive values were calculated based on the theorem of Bayes using Fagan's nomogram. RESULTS: Of 165 participants (56.4% male, 71 ± 16 years) included in the study, the diagnosis of COVID-19 was confirmed with RT-PCR and AB tests in 13 participants (prevalence 7.9%). Sensitivity and specificity of chest CT were 84.6% (95% confidence interval [CI], 54.6-98.1) and 94.7% (95% CI, 89.9-97.7), respectively. Positive and negative likelihood ratio of chest CT were 16.1 (95% CI, 7.9-32.8) and 0.16 (95% CI, 0.05-0.58) and positive and negative predictive value were 57.9% (95% CI, 40.3-73.7) and 98.6% (95% CI, 95.3-99.6), respectively. CONCLUSION: At a low prevalence of COVID-19, chest CT could be used as a complement to repeated RT-PCR testing for early COVID-19 exclusion in adults with suspected infection before referral to hospital's general wards. Trial registration ClinicalTrials.gov: NCT04357938 April 22, 2020.

Exotoxins from Staphylococcus aureus activate 5-lipoxygenase and induce leukotriene biosynthesis.

Massive neutrophil infiltration is an early key event in infectious inflammation, accompanied by chemotactic leukotriene (LT)B4 generation. LTB4 biosynthesis is mediated by 5-lipoxygenase (5-LOX), but which pathogenic factors cause 5-LOX activation during bacterial infections is elusive. Here, we reveal staphylococcal exotoxins as 5-LOX activators. Conditioned medium of wild-type Staphylococcus aureus but not of exotoxin-deficient strains induced 5-LOX activation in transfected HEK293 cells. Two different staphylococcal exotoxins mimicked the effects of S. aureus-conditioned medium: (1) the pore-forming toxin α-hemolysin and (2) amphipathic α-helical phenol-soluble modulin (PSM) peptides. Interestingly, in human neutrophils, 5-LOX activation was exclusively evoked by PSMs, which was prevented by the selective FPR2/ALX receptor antagonist WRW4. 5-LOX activation by PSMs was confirmed in vivo as LT formation in infected paws of mice was impaired in response to PSM-deficient S. aureus. Conclusively, exotoxins from S. aureus are potent pathogenic factors that activate 5-LOX and induce LT formation in neutrophils.

Heater-Cooler Devices and Risk of Contamination during Cardiac Surgery.

BACKGROUND: Heater-cooler devices (HCD) have been implicated in a cardiosurgical contamination scenario causing prosthetic valve endocarditis. AIM: We characterized contamination of new HCDs and assessed the risk of intraoperative microorganism transmission from the HCD to the operating field. METHODS: We initially acquired four new FlexTherm and then four new Maquet HCU40 HCDs and assessed occurrence and speed of microbial contamination (including mycobacteria) assessing swab and water samples from the device. In parallel, we collected repeated samples from different sites in the operating room either by swab sticks or by exposing different sample plates to room air. We also reviewed microbiological results from the hospital and compared them to cardiosurgical wound infections and endocarditis cases. Finally, we simulated cardiosurgical conditions and assessed the devices' ability to expel air to the operative field. RESULTS: All new HCDs were clean before first use. Despite authority-mandated decontamination procedures, microbial growth (Fusarium solani, Sphingomonas paucimobilis, Pseudomonas aeruginosa, Mycobacterium chelonae, and gordonae) was identified in all HCDs over time and could not be permanently eliminated. Four of these mircoorganisms were also found in tap water. However, none of the HCD-organisms were found inside the laminar airflow operating area. Importantly, except for P. aeruginosa, none of the HCD organisms were found in patients with surgical wound infections or endocarditis. HCD-expelled air did not rise more than 40 cm above ground. CONCLUSION: HCDs cannot be expected to remain sterile despite extensive decontamination procedures. However, airborne transmission of microorganisms directly from the HCD to the operating field appears unlikely.

The Inflammatory Profile of Obesity and the Role on Pulmonary Bacterial and Viral Infections.

Obesity is a globally increasing health problem, entailing diverse comorbidities such as infectious diseases. An obese weight status has marked effects on lung function that can be attributed to mechanical dysfunctions. Moreover, the alterations of adipocyte-derived signal mediators strongly influence the regulation of inflammation, resulting in chronic low-grade inflammation. Our review summarizes the known effects regarding pulmonary bacterial and viral infections. For this, we discuss model systems that allow mechanistic investigation of the interplay between obesity and lung infections. Overall, obesity gives rise to a higher susceptibility to infectious pathogens, but the pathogenetic process is not clearly defined. Whereas, viral infections often show a more severe course in obese patients, the same patients seem to have a survival benefit during bacterial infections. In particular, we summarize the main mechanical impairments in the pulmonary tract caused by obesity. Moreover, we outline the main secretory changes within the expanded adipose tissue mass, resulting in chronic low-grade inflammation. Finally, we connect these altered host factors to the influence of obesity on the development of lung infection by summarizing observations from clinical and experimental data.

Diagnostic accuracy and clinical impact of loop-mediated isothermal amplification for rapid detection of Staphylococcus aureus bacteremia: a retrospective observational study.

The aim of this study was to evaluate the diagnostic accuracy and the clinical impact of isothermal loop-mediated amplification (LAMP; eazyplex® MRSA kits) for rapid diagnosis of Staphylococcus aureus bacteremia (SAB) in comparison with conventional blood culture diagnostics. We performed a retrospective, single-center observational study over the period between November 2016 and December 2018 on patients (and blood cultures) with growth of Gram-positive cocci in clusters in their blood cultures. We quantified diagnostic accuracy with sensitivity and specificity for detection of S. aureus, methicillin-resistant S. aureus (MRSA), and the mecA/C resistance genes in 797 blood cultures. The clinical impact was assessed by time to result reporting, time to appropriate treatment, and length of stay in intensive care unit (ICU) in 190 SAB patients. We observed sensitivity and specificity above 90% for S. aureus detection (sensitivity (95% confidence interval (CI)), 99.57% (97.61%, 99.98%); specificity, 99.12% (97.95%, 99.71%)), for MRSA detection (sensitivity, 100% (89.11%, 100.00%); specificity, 99.72% (99.05, 99.96)), and for mecA/C detection (sensitivity, 94.71% (91.85%, 96.78%); specificity, 95.89% (93.58%, 97.54%)). LAMP testing was associated with shorter median time to result reporting (24.0 h (first and third quartiles (Q1-Q3), 20.0-27.0 h) vs 41.5 h (36.0-46.0 h); p < 0.001) and different distribution of time to appropriate treatment (2.0 days (1.0-3.0) vs 2.0 days (2.0-3.0); p = 0.004). No evidence for differences in length-of-stay in ICU was observed. Our analysis suggests for the application of LAMP (i) a high diagnostic accuracy for detection of S. aureus and the mecA/C genes in blood cultures, (ii) an earlier result reporting, and (iii) a shorter time to appropriate treatment.

Rapid culture-based identification of Shiga toxin-producing Escherichia coli and Shigella spp./Enteroinvasive E. coli using the eazyplex® EHEC complete assay.

Shiga toxin-producing Escherichia coli (STEC) and Shigella spp./enteroinvasive E. coli (EIEC) are common diarrheagenic bacteria that cause sporadic diseases and outbreaks. Clinical manifestations vary from mild symptoms to severe complications. For microbiological diagnosis, culture confirmation of a positive stool screening PCR test is challenging because of time-consuming methods for isolation of strains, wide variety of STEC pathotypes, and increased emergence of non-classical strains with unusual serotypes. Therefore, molecular assays for the rapid identification of suspect colonies growing on selective media are very useful. In this study, the performance of the newly introduced eazyplex® EHEC assay based on loop-mediated isothermal amplification (LAMP) was evaluated using 18 representative STEC and Shigella strains and 31 isolates or positive-enrichment broths that were collected from clinical stool samples following screening by BD MAX™ EBP PCR. Results were compared to real-time PCR as a reference standard. Overall, sensitivities and specificities of the eazyplex® EHEC were as follows: 94.7% and 100% for Shiga toxin 1 (stx1), 100% and 100% for stx2, 93.3% and 97.1% for intimin (eae), 100% and 100% for enterohemolysin A (ehlyA), and 100% and 100% for invasion-associated plasmid antigen H (ipaH) as Shigella spp./EIEC target, respectively. Sample preparation for LAMP took only some minutes, and the time to result of the assay ranged from 8.5 to 13 min. This study shows that eazyplex® EHEC is a very fast and easy to perform molecular assay that provides reliable results as a culture confirmation assay for the diagnosis of STEC and Shigella spp./EIEC infections.