Antibiotics, Vol. 8, Pages 85: Antimicrobial Resistance in Bacterial Pathogens and Detection of Carbapenemases in Klebsiella pneumoniae Isolates from Hospital Wastewater During a six-month period (October 2017–March 2018), the prevalence and susceptibility of important pathogenic bacteria isolated from 12 hospital raw sewage samples in North Western Greece was investigated. The samples were analyzed for methicillin-resistant Staphylococcus aureus (MRSA), vancomycin-resistant enterococci (VRE), extended-spectrum beta-lactamase (ESBL) producing Escherichia coli, carbapenemase-producing Klebsiella pneumoniae (CKP), and multidrug-resistant Pseudomonas aeruginosa. Antimicrobial susceptibility testing was performed using the agar diffusion method according to the recommendations of the Clinical and Laboratory Standards Institute. The diversity of carbapenemases harboring K. pneumoniae was examined by two phenotyping screening methods (modified Hodge test and combined disk test), a new immunochromatographic rapid assay (RESIST-4 O.K.N.V.) and a polymerase chain reaction (PCR). The results demonstrated the prevalence of MRSA, vancomycin-resistant Staphylococcus aureus (VRSA), VRE, and CKP in the examined hospital raw sewage samples. In addition, the aforementioned methods which are currently used in clinical laboratories for the rapid identification and detection of resistant bacteria and genes, performed sufficiently to provide reliable results in terms of accuracy and efficiency.

Antibiotics, Vol. 8, Pages 84: Qualitative Analysis of Primary Care Provider Prescribing Decisions for Urinary Tract Infections Inappropriate choices and durations of therapy for urinary tract infections (UTI) are a common and widespread problem. In this qualitative study, we sought to understand why primary care providers (PCPs) choose certain antibiotics or durations of treatment and the sources of information they rely upon to guide antibiotic-prescribing decisions. We conducted semi-structured interviews with 18 PCPs in two family medicine clinics focused on antibiotic-prescribing decisions for UTIs. Our interview guide focused on awareness and familiarity with guidelines (knowledge), acceptance and outcome expectancy (attitudes), and external barriers. We followed a six-phase approach to thematic analysis, finding that many PCPs believe that fluoroquinolones achieve more a rapid and effective control of UTI symptoms than trimethoprim-sulfamethoxazole or nitrofurantoin. Most providers were unfamiliar with fosfomycin as a possible first-line agent for the treatment of acute cystitis. PCPs may be misled by advanced patient age, diabetes, and recurrent UTIs to make inappropriate choices for the treatment of acute cystitis. For support in clinical decision making, few providers relied on guidelines, preferring instead to have decision support embedded in the electronic medical record. Knowing the PCPs’ knowledge gaps and preferred sources of information will guide the development of a primary care-specific antibiotic stewardship intervention for acute cystitis.

Antibiotics, Vol. 8, Pages 83: Dissolution of the Disparate: Co-ordinate Regulation in Antibiotic Biosynthesis Discovering new antibiotics is vital to combat the growing threat of antimicrobial resistance. Most currently used antibiotics originate from the natural products of actinomycete bacteria, particularly Streptomyces species, that were discovered over 60 years ago. However, genome sequencing has revealed that most antibiotic-producing microorganisms encode many more natural products than previously thought. Biosynthesis of these natural products is tightly regulated by global and cluster situated regulators (CSRs), most of which respond to unknown environmental stimuli, and this likely explains why many biosynthetic gene clusters (BGCs) are not expressed under laboratory conditions. One approach towards novel natural product discovery is to awaken these cryptic BGCs by re-wiring the regulatory control mechanism(s). Most CSRs bind intergenic regions of DNA in their own BGC to control compound biosynthesis, but some CSRs can control the biosynthesis of multiple natural products by binding to several different BGCs. These cross-cluster regulators present an opportunity for natural product discovery, as the expression of multiple BGCs can be affected through the manipulation of a single regulator. This review describes examples of these different mechanisms, including specific examples of cross-cluster regulation, and assesses the impact that this knowledge may have on the discovery of novel natural products.

Antibiotics, Vol. 8, Pages 82: A Topical Desiccant Agent in Association with Manual Debridement in the Initial Treatment of Peri-Implant Mucositis: A Clinical and Microbiological Pilot Study In patients presenting mucositis, effective sub-gingival debridement is crucial to prevent peri-implantitis. The aim of this randomized study was to assess the three-month (T1) effects of a locally delivered liquid desiccant agent with molecular hygroscopic properties, in association with manual debridement, at sites with peri-implant mucositis. Twenty-three patients presenting at least one implant with no radiographically detectable bone loss, a pocket probing depth (PPD) ≥ 4 mm, and bleeding on probing (BOP), were included. At baseline (T0), patients were randomly assigned to receive the aforementioned desiccant agent before debridement (Test-Group), or a Chlorhexidine 1% disinfectant gel after debridement (Control-Group). Treatments were repeated after seven and 14 days. Peri-implant soft tissue assessment [PPD, BOP, Modified Bleeding Index (mBI), Visible Plaque Index (VPI), and Modified Plaque Index (mPLI)] and microbial sampling were performed at T0 and T1. At T1 the Test-Group presented significantly greater reductions for BOP, mBI, VPI, and mPLI. Concerning the deepest sites of the treated implants, both groups showed statistically significant reductions for BOP and mBI between T0 and T1. Furthermore, the Test-Group exhibited a significant decrease in anaerobic bacteria. Despite these valid outcomes, a complete resolution of the inflammatory conditions was not achieved by any of the groups.

Antibiotics, Vol. 8, Pages 81: Impact of Mobility on Methicillin-Resistant Staphylococcus aureus among Injection Drug Users In this study, we develop and present a deterministic model for the transmission dynamics of methicillin-resistant staphylococcus aureus (MRSA) among injection drug users. The model consists of non-injection drug users as well as low-and high-risk injection drug users (IDUs). The model further incorporates the movement of these individuals between large metro, suburban and rural areas. The model parameters were estimated by fitting the model to the 2008–2013 disease prevalence data for non-IDUs obtained from the Agency for Healthcare and Research and Quality (AHRQ), as well as the 2009–2013 Census Bureau data for the number of individuals migrating between three different counties in Kansas. Sensitivity analysis was implemented to determine the parameters with the most significant impact on the total number of infected individuals; the transmission probability, recovery rates, and positive behavioral change parameter for the subgroup have the most significant effect on the number of infected individuals. Furthermore, the sensitivity of the parameters in the different areas was the same when the areas are disconnected. When the areas are connected, the parameters in large-metro areas were the most sensitive, and the rural areas were least sensitive. The result shows that to effectively control the disease across the large metro, suburban and rural areas, it is best to focus on controlling both behavior and disease in the large metro area as this has a trickle-down effect to the other places. However, controlling behavior and disease at the same time in all the areas will lead to the elimination of the disease.

Antibiotics, Vol. 8, Pages 80: Complement Evasion in Borrelia spirochetes: Mechanisms and Opportunities for Intervention Lyme disease (LD) is an increasingly prevalent, climate change-accelerated, vector-borne infectious disease with significant morbidity and cost in a proportion of patients who experience ongoing symptoms after antibiotic treatment, a condition known as post-treatment Lyme disease syndrome (PTLDS). Spirochetal bacteria of Borrelia species are the causative agents of LD. These obligate parasites have evolved sophisticated immune evasion mechanisms, including the ability to defeat the innate immune system’s complement cascade. Research on complement function and Borrelia evasion mechanisms, focusing on human disease, is reviewed, highlighting opportunities to build on existing knowledge. Implications for the development of new antibiotic therapies having the potential to prevent or cure PTLDS are discussed. It is noted that a therapy enabling the complement system to effectively counter Borrelia might have lower cost and fewer side-effects and risks than broad-spectrum antibiotic use and could avert the need to develop and administer a vaccine.

Antibiotics, Vol. 8, Pages 79: Antibiotic Prescribing Quality in Out-of-Hours Primary Care and Critical Appraisal of Disease-Specific Quality Indicators Outpatient antibiotic use in Belgium is among the highest in Europe. The most common reason for an encounter in out-of-hours (OOH) primary care is an infection. In this study, we assessed all consultations from July 2016 to June 2018 at five OOH services. We described antibiotic prescribing by diagnosis, calculated disease-specific antibiotic prescribing quality indicators’ (APQI) values and critically appraised these APQI. We determined that 111,600 encounters resulted in 26,436 (23.7%) antibiotic prescriptions. The APQI diagnoses (i.e., bronchitis, upper respiratory infection, cystitis, tonsillitis, sinusitis, otitis media, and pneumonia) covered 14,927 (56.7%) antibiotic prescriptions. Erysipelas (1344 (5.1%)) and teeth/gum disease (982 (3.7%)) covered more prescriptions than sinusitis or pneumonia. Over 75% of patients with tonsillitis and over 50% with bronchitis, sinusitis, and otitis media were prescribed an antibiotic. Only for otitis media the choice of antibiotic was near the acceptable range. Over 10% of patients with bronchitis or pneumonia and over 25% of female patients with an acute cystitis received quinolones. The APQI cover the diagnoses for only 57% of all antibiotic prescriptions. As 5.1% and 3.7% of antibiotic prescriptions are made for erysipelas and teeth/gum disease, respectively, we propose to add these indications when assessing antibiotic prescribing quality in OOH primary care.

Antibiotics, Vol. 8, Pages 78: Levofloxacin-Associated Neurotoxicity in a Patient with a High Concentration of Levofloxacin in the Blood and Cerebrospinal Fluid Neurotoxicity is a rare and intolerable adverse effect associated with levofloxacin therapy, whose diagnosis has mostly been reported based on medical history rather than quantitative measures in the blood. We report a 68-year-old man with levofloxacin-associated encephalopathy and myoclonus with high levels of levofloxacin in the blood and cerebrospinal fluid. After hemodialysis, these decreased, and his symptoms rapidly improved. An electroencephalogram was also normal. This case showed the concentration of levofloxacin to be clearly related to levofloxacin-associated neurotoxicity. Therefore, an estimation of its concentration may contribute to accurate diagnosis.

Antibiotics, Vol. 8, Pages 77: Inhibition of Bacterial and Fungal Biofilm Formation by 675 Extracts from Microalgae and Cyanobacteria Bacterial biofilms are complex biological systems that are difficult to eradicate at a medical, industrial, or environmental level. Biofilms confer bacteria protection against external factors and antimicrobial treatments. Taking into account that about 80% of human infections are caused by bacterial biofilms, the eradication of these structures is a great priority. Biofilms are resistant to old-generation antibiotics, which has led to the search for new antimicrobials from different sources, including deep oceans/seas. In this study, 675 extracts obtained from 225 cyanobacteria and microalgae species (11 phyla and 6 samples belonging to unknown group) were obtained from different culture collections: The Blue Biotechnology and Ecotoxicology Culture Collection (LEGE-CC), the Coimbra Collection of Algae (ACOI) from Portugal, and the Roscoff Culture Collection (RCC) from France. The largest number of samples was made up of the microalgae phylum Chlorophyta (270) followed by Cyanobacteria (261). To obtain a large range of new bioactive compounds, a method involving three consecutive extractions (hexane, ethyl acetate, and methanol) was used. The antibiofilm activity of extracts was determined against seven different bacterial species and two Candida strains in terms of minimal biofilm inhibitory concentration (MBIC). The highest biofilm inhibition rates (%) were achieved against Candida albicans and Enterobacter cloacae. Charophyta, Chlorophyta, and Cyanobacteria were the most effective against all microorganisms. In particular, extracts of Cercozoa phylum presented the lowest MBIC50 and MBIC90 values for all the strains except C. albicans.

Antibiotics, Vol. 8, Pages 76: The Link between Purine Metabolism and Production of Antibiotics in Streptomyces Stress and starvation causes bacterial cells to activate the stringent response. This results in down-regulation of energy-requiring processes related to growth, as well as an upregulation of genes associated with survival and stress responses. Guanosine tetra- and pentaphosphates (collectively referred to as (p)ppGpp) are critical for this process. In Gram-positive bacteria, a main function of (p)ppGpp is to limit cellular levels of GTP, one consequence of which is reduced transcription of genes that require GTP as the initiating nucleotide, such as rRNA genes. In Streptomycetes, the stringent response is also linked to complex morphological differentiation and to production of secondary metabolites, including antibiotics. These processes are also influenced by the second messenger c-di-GMP. Since GTP is a substrate for both (p)ppGpp and c-di-GMP, a finely tuned regulation of cellular GTP levels is required to ensure adequate synthesis of these guanosine derivatives. Here, we discuss mechanisms that operate to control guanosine metabolism and how they impinge on the production of antibiotics in Streptomyces species.