Human Tularemia Epididymo-Orchitis Caused by Francisella tularensis Subspecies holartica, Austria.

A previously healthy man in Austria had tularemia epididymo-orchitis develop, leading to unilateral orchiectomy. Francisella tularensis subspecies holartica was detected by 16S rRNA gene sequencing analysis of inflamed granulomatous testicular tissue. Clinicians should suspect F. tularensis as a rare etiologic microorganism in epididymo-orchitis patients with relevant risk factors.

TLR4/CD14/MD2 Revealed as the Limited Toll-like Receptor Complex for Chlamydia trachomatis-Induced NF-κB Signaling.

Chlamydia trachomatis (Ct) is the most common cause of genital tract infections as well as preventable blindness worldwide. Pattern recognition receptors such as toll-like receptors (TLRs) represent the initial step in recognizing pathogenic microorganisms and are crucial for the initiation of an appropriate immune response. However, our understanding of TLR-signaling in Chlamydia-infected immune cells is incomplete. For a better comprehension of pathological inflammatory responses, robust models for interrogating TLR-signaling upon chlamydial infections are needed. To analyze the TLR response, we developed and utilized a highly sensitive and selective fluorescent transcriptional cellular reporter system to measure the activity of the transcription factor NF-κB. Upon incubation of the reporter cells with different preparations of Ct, we were able to pinpoint which components of TLRs are involved in the recognition of Ct. We identified CD14 associated with unique characteristics of different serovars as the crucial factor of the TLR4/CD14/MD2 complex for Ct-mediated activation of the NF-κB pathway. Furthermore, we found the TLR4/CD14/MD2 complex to be decisive for the uptake of Ct-derived lipopolysaccharides but not for infection and replication of Ct. Imaging flow cytometry provided information about inclusion formation in myeloid- as well as lymphocytic cells and was highest for Ct L2 with at least 25% of inclusion forming cells. Ct E inclusion formation was eminent in Jurkat cells without CD14 expression (11.1%). Thus, our model enables to determine Ct uptake and signal induction by pinpointing individual components of the recognition and signaling pathways to better understand the immune response towards infectious pathogens.

Tick feeding modulates the human skin immune landscape to facilitate tick-borne pathogen transmission.

During cutaneous tick attachment, the feeding cavity becomes a site of transmission for tick salivary compounds and tick-borne pathogens. However, the immunological consequences of tick feeding for human skin remain unclear. Here, we assessed human skin and blood samples upon tick bite and developed a human skin explant model mimicking Ixodes ricinus bites and tick-borne pathogen infection. Following tick attachment, we observed rapidly occurring patterns of immunomodulation, including increases in neutrophils and cutaneous B and T cells. T cells upregulated tissue residency markers, while lymphocytic cytokine production was impaired. In early stages of Borrelia burgdorferi model infections, we detected strain-specific immune responses and close spatial relationships between macrophages and spirochetes. Preincubation of spirochetes with tick salivary gland extracts hampered accumulation of immune cells and increased spirochete loads. Collectively, we showed that tick feeding exerts profound changes on the skin immune network that interfere with the primary response against tick-borne pathogens.

A Novel Flow Cytometric Approach for the Quantification and Quality Control of Chlamydia trachomatis Preparations.

Chlamydia trachomatis is an obligate intracellular pathogenic bacterium with a biphasic developmental cycle manifesting two distinct morphological forms: infectious elementary bodies (EBs) and replicative intracellular reticulate bodies (RBs). Current standard protocols for quantification of the isolates assess infectious particles by titering inclusion-forming units, using permissive cell lines, and analyzing via immunofluorescence. Enumeration of total particle counts is achieved by counting labeled EBs/RBs using a fluorescence microscope. Both methods are time-consuming with a high risk of observer bias. For a better assessment of C. trachomatis preparations, we developed a simple and time-saving flow cytometry-based workflow for quantifying small particles, such as EBs with a size of 300 nm. This included optimization of gain and threshold settings with the addition of a neutral density filter for small-particle discrimination. The nucleic acid dye SYBR® Green I (SGI) was used together with propidium iodide and 5(6)-carboxyfluorescein diacetate to enumerate and discriminate between live and dead bacteria. We found no significant differences between the direct particle count of SGI-stained C. trachomatis preparations measured by microscopy or flow cytometry (p > 0.05). Furthermore, we completed our results by introducing a cell culture-independent viability assay. Our measurements showed very good reproducibility and comparability to the existing state-of-the-art methods, indicating that the evaluation of C. trachomatis preparations by flow cytometry is a fast and reliable method. Thus, our method facilitates an improved assessment of the quality of C. trachomatis preparations for downstream applications.

The association between shift patterns and the quality of hand antisepsis in a neonatal intensive care unit: An observational study.

BACKGROUND: Healthcare-associated infections represent a major burden in neonatal intensive care units. Hand antisepsis is the most important tool for prevention, however, compliance among healthcare workers remains low. OBJECTIVES: To prospectively evaluate the influence of different work shifts (extended working hours, night shifts) on the quality of healthcare workers' hand antisepsis. DESIGN: Observational study. SETTINGS: Two equivalent "Level III" neonatal intensive care units at the University Hospital Vienna, Austria. PARTICIPANTS: Seventy healthcare workers, 46 nurses and 24 physicians. METHODS: The Semmelweis Scanner, an innovative training device assessing the quality of hand antisepsis with an ultraviolet dye labelled alcohol-based hand rub, was employed to collect data on the hand surface coverage achieved during hand antisepsis of participants. It provides visual feedback of appropriately versus inappropriately disinfected areas of the hand and can also be used for the objective quantification of hand surface coverage with the hand rub. Measurements were performed before and after 12.5 h (h) day and night shifts (nurses), as well as before and after regular 8 h day shifts and extended 25 h shifts (physicians). To avoid any bias caused by residual ultraviolet marker, scans had to be separated by 24 h periods. Primary outcome was the hand surface coverage with the hand rub: Hand scans were categorized as "passed" if an appropriate quality of hand hygiene, defined as a minimum 97% coverage of hand surface, was achieved. A generalized mixed model was used to analyse the data accounting for repeated measurements. RESULTS: Seventy healthcare workers performed a total of 485 scans. Nineteen scans had to be excluded, resulting in 466 scans for further analyses. A difference in the predicted probability of achieving appropriate hand antisepsis was found between the beginning and end of extended shifts: In physicians, adequate hand antisepsis was remarkably reduced after 25 h shifts (predicted probability 99.4% vs 78.8%), whereas there was no relevant difference between the beginning and end of 8 h day shifts (92.2% vs 97.3%). In nurses, a relevant difference was found between the beginning and end of 12.5 h day shifts (88.6% vs 73.6%). This difference was not found for 12.5 h night shifts. The most frequently missed area on the hands was the right dorsum. CONCLUSION: The quality of hand antisepsis of healthcare workers in neonatal intensive care units may be associated with long working hours.

Invasive Fungal Infection Caused by Exophiala dermatitidis in a Patient After Lung Transplantation: Case Report and Literature Review.

This report describes a case of invasive Exophiala dermatitidis infection after double lung transplantation in a 76-year-old man. After thoracotomy, the patient's wound showed dehiscence with purulent secretion. The black yeast was isolated from cultures taken from the wound, and species identification was confirmed by sequence analysis of the internal transcribed spacer (ITS-S2) region. The results of the susceptibility testing showed voriconazole as the most active drug. Despite adaptation of the antifungal therapy the clinical condition worsened, and the patient died. In addition, we evaluated the fungicidal activity of antiseptics towards E. dermatitidis and aimed to provide a brief literature review of previously reported infections caused by this rare fungus. To the best of our knowledge, this is the first report of a rapidly progressing invasive fungal infection with E. dermatitidis originating from a colonized wound after lung transplantation.

Epidemiology of Urinary Tract Infections in HIV Positive Patients at a Tertiary Care University Hospital in Central Europe (2011 - 2016).

OBJECTIVES: There is limited published data concerning the recent epidemiology of urinary tract infections (UTI) in HIV-patients, thus we analysed independent risk factors for UTI in HIV positive individuals and antimicrobial resistance rates of E. coli to antimicrobial agents commonly used in UTI. To determine the prevalence of symptomatic urinary tract infections (UTI) in HIV-patients, we performed a retrospective case-control study. METHODS: We included 313 HIV-patients, 101 with UTI and 212 age and gendermatched controls, attending the HIV outpatient clinic at the Vienna University Hospital (VUH) over a period from January 2011 to September 2016. The patients' specific data was gathered from the electronic database of the VUH. The statistical analysis was performed using SPSS Software Version 20.0. RESULTS: HIV infected individuals with CD4 count >200 cells/mm3 were less likely than HIV infected individuals with CD4 count <200 cells/mm3 to experience UTI (OR 0.811, 95% CI 0.712-0.923 vs. OR 2.555, 95% CI 1.553 - 4.205, respectively). The in vitro resistance rate of E. coli to antimicrobial agents was as follows: ciprofloxacin (41%), mecillinam (20.5%), trimethoprim (61%), ampicillin (67%), ampicillin/ clavulanic acid (23%), cefuroxime (17%), nitrofurantoin (2%), amikacin (0%) and gentamicin (9.5%). CONCLUSION: Immunological status (CD4 count) is an important parameter for risk assessment of UTIs in HIV-patients. The increased resistance rate of E. coli to commonly used antimicrobial agents needs to be considered when it comes to the management of UTI, additionally, surveillance strategies should be implemented in HIV-patients.

Clinical characteristics of a large cohort of patients with positive culture of Fusobacterium necrophorum.

Background: Fusobacterium necrophorum is a rare pathogen, mostly affecting young adults, causing infections of the head and neck, typically described as the Lemierre's syndrome. Today this symptom complex has become increasingly rare and has almost turned to a 'forgotten disease'. Methods: We performed a retrospective, descriptive study to identify the clinical features of patients with positive culture of F. necrophorum. Additionally, the antibiotic susceptibility profile of the pathogens was analysed. Results: During a period of 22 years 36 patients with at least one isolate of F. necrophorum were identified. Mostly tonsillar and peritonsillar abscesses were found, 10 patients were identified with bacteraemia, but only 4 patients presented with symptoms like sore throat, fever and swollen cervical lymph nodes, which may suggest Lemierre's. Most of the isolates (33/35) showed sensitivity to all tested antibiotics. Conclusion: Appropriate techniques are needed to detect F. necropho rum, especially from throat swabs, in the microbiological laboratory. Current clinical and microbiological practice may lead to under-diagnosis of infections caused by F. necrophorum. Further research is needed to define the colonization rate and to optimize methods for detection as well as identification of virulence.

Evaluation of three experimental in vitro models for the assessment of the mechanical cleansing efficacy of wound irrigation solutions.

The aim of this study was to compare different wound-rinsing solutions to determine differences in the efficiency and to evaluate three different in vitro models for wound cleansing. Different wound-rinsing solutions (physiological saline solution, ringer lactate solution for wound irrigation, water and a solution containing polihexanide and the surfactant undecylenamidopropyl-betain) were applied on standardised test models (one- and three-chamber model, flow-cell method and a biofilm model), each challenged with three different standardised wound test soils. In the one-chamber model saline showed a better effect on decontaminating proteins than the ringer lactate solution. In the flow-cell method, water performed better than physiological saline solution, whereas ringer lactate solution demonstrated the lowest cleansing effect. No obvious superiority between the two electrolyte-containing solutions was detectable in the biofilm model. Unfortunately, it was not possible to assess the protein decontamination qualities of the surfactant-containing solution because of the interference with the protein measurement. The flow-cell method was able to detect differences between different rinse solutions because it works at constant flow mechanics, imitating a wound-rinsing procedure. The three-chamber and the less-pronounced modified one-chamber method as well as the biofilm model had generated inhomogeneous results.

Shortening the Application Time of Alcohol-Based Hand Rubs to 15 Seconds May Improve the Frequency of Hand Antisepsis Actions in a Neonatal Intensive Care Unit.

BACKGROUND For alcohol-based hand rubs, the currently recommended application time of 30 seconds is longer than the actual time spent in clinical practice. We investigated whether a shorter application time of 15 seconds is microbiologically safe in neonatal intensive care and may positively influence compliance with the frequency of hand antisepsis actions. METHODS We conducted in vitro experiments to determine the antimicrobial efficacy of hand rubs within 15 seconds, followed by clinical observations to assess the effect of a shortened hand antisepsis procedure under clinical conditions in a neonatal intensive care unit (NICU). An independent observer monitored the frequency of hand antisepsis actions during shifts. RESULTS All tested hand rubs fulfilled the requirement of equal or even significantly higher efficacy within 15 seconds when compared to a reference alcohol propan-2-ol 60% (v/v) within 30 seconds. Microbiologically, reducing the application time to 15 seconds had a similar effect when compared to 30-second hand rubbing, but it resulted in significantly increased frequency of hand antisepsis actions (7.9±4.3 per hour vs 5.8±2.9 per hour; P=.05). CONCLUSION Time pressure and workload are recognized barriers to compliance. Therefore, reducing the recommended time for hand antisepsis actions, using tested and well-evaluated hand rub formulations, may improve hand hygiene compliance in clinical practice. Infect Control Hosp Epidemiol 2017;38:1430-1434.