Current methods for microbiological diagnosis of acute central nervous system infections

The incidence of infections affecting the central nervous system has increased in recent years, making neuroinfections a current global health problem. The central nervous system is quite well protected from the external and internal environments, although it is susceptible to infection by a wide variety of pathogens. The etiological diversity further complicates the management of such infections because it is important to identify correctly the specific cause in order to choose the most appropriate antimicrobial therapy. Diagnosis is made not only based on clinical and epidemiological data but also on the results of clinical laboratory and microbiological examination of cerebrospinal fluid. This article aims to review current microbiological methods in the diagnosis of acute central nervous system infections and help healthcare providers to recognize their advantages and limitations in order to manage their patients appropriately

The Programmed Cell Death of an Immature Thymocyte Cell Line Transgenic for an αβ TCR and the c-myc Proto-Oncogene

The c-myc proto-oncogene linked to the mouse Thy-1 gene transcriptional unit predisposes mice to development of thymic tumors consisting predominantly of immature CD4+ CD8+ cells. In an attempt to immortalize immature T cells expressing a known T-cell antigen receptor (TCR), Thy-1/c-myc transgenic mice were bred to αβ TCR transgenic mice (F5), and CD4+ CD8+ cell lines were established from thymic tumors in double-transgenic mice. These cells expressed high-level heat-stable antigen (HSA) and were able to undergo programmed cell death upon induction with steroids and CD3 cross-linking, but not with cognate peptide. In addition, one line had rearranged and transcribed endogenous TCR c and genes, in spite of the fact that transgenic α and β genes were also expressed. Furthermore, we show that Thy-1/myc transgenic mice deficient in recombination activating gene-1 (RAG-1) do not develop tumors, in contrast to RAG-1-/- mice, which are also transgenic for both Thy-1/myc and the F5 TCR. This indicates that in order for thymocytes to be transformed by the Thy-myc transgene, they need to proceed to the double-positive stage

MCCOY AND MCCOY-PLOVDIV CELL LINES IN EXPERIMENTAL AND DIAGNOSTIC PRACTICE - PAST, PRESENT AND PERSPECTIVES

The McCoy cell line has almost 50 years history. The cells are widely applied in the diagnostics and culture of various microorganisms with medical importance. The cell line is included in laboratory and diagnostic tests which are the basis for study of interactions between various pathogens and host cells leading to cytotoxic damage or cell death. With its importance in experimental and diagnostic laboratories, McCoy cell line is among the most popular cell cultures - HeLa, HEp-2, Vero, CaCo-2, 3T3, MDCK. An alternative for application of McCoy is the serum-free strain McCoy-Plovdiv. It is cultured in completely defined, serum- and protein-free medium. It keeps the properties of the parental line but also offers new opportunities

Tolerance in TCR/Cognate Antigen Double-Transgenic Mice Mediated by Incomplete Thymic Deletion and Peripheral Receptor Downregulation

Influenza nucleoprotein (NP)-specific T-cell receptor transgenic mice (F5) were crossed with transgenic mice expressing the cognate antigenic protein under the control of the H- 2Kb promoter. Double-transgenic mice show negative selection of thymocytes at the CD4+8+TCR10 to CD4+8+TCRhi transition stage. A few CD8 T cells, however, escape clonal deletion, and in the peripheral lymphoid organs of these mice, they exhibit low levels of the transgenic receptor and upregulated levels of the CD44 memory marker. Such cells do not proliferate upon exposure to antigen stimulation in vivo or ex vivo, however, they can develop low but detectable levels of antigen-specific cytotoxic function after stimulation in vitro in the presence of IL-2

Vitamin D3 exerts immunomodulatory and memory improving properties in rats with lipopolysaccharide-induced inflammation

Introduction: Vitamin D is a fat-soluble secosteroid, its primary function being regulation of calcium-phosphate homeostasis and maintenance of bone integrity and mineralization. Recently, pleotropic effects of this vitamin have been recognized, including an immunomodulatory role and involvement in normal brain development and functioning. Aim: The aim of the present study was to investigate the influence of cholecalciferol on serum inflammatory markers and memory functions in lipopolysaccharide (LPS) model of inflammation. Materials and methods: Male Wistar rats were randomly divided into 4 groups (n=8): control group, LPS control group, LPS + cholecalciferol (vitamin D3) 500 UI group, and 1000 IU/kg bw group. Step-down passive avoidance test, novel object recognition test (NORT), Y- and T-maze were performed to assess the memory functions. Latency, recognition index (RI), % spontaneous alteration (SA), and working memory index were registered. Tumor necrosis factor-alpha (TNF-α), IL-1β, transforming growth factor-β1 (TGF-β1), and brain derived neurotrophic factor (BDNF) serum levels were measured by ELISA. Results: LPS administration caused significant impairment in memory functions in all memory tasks. Cholecalciferol treatment caused significant increase in % SA, RI, and working memory index. In the step-down passive avoidance test, cholecalciferol-treated groups showed statistically significant increase in latency in the long-term memory test. Vitamin D3-treated rats showed decreased TNF-α and IL-1β serum levels whereas the concentration of TGF-β1 and BDNF increased. Conclusions: Cholecalciferol improves spatial working and episodic memory, which can at least partially be explained with its effect on systemic inflammatory response that is closely related with the development of neuroinflammation

ASSESSMENT OF SARS-COV-2 SPECIFIC B-CELL IMMUNE MEMORY: EVIDENCE FOR PERSISTENCE UP TO 1 YEAR POST-INFECTION

Background: SARS-CoV-2, the virus responsible for COVID-19 pandemic, has posed huge global health challenges. Understanding the immune response to SARS-CoV-2 infection, and in particular – the role of B cells in the generation of immune memory is crucial for assessing the durability of protective immunity. Materials and Methods: In this longitudinal prospective study, individuals who had recovered from SARS-CoV-2 infection were included. Peripheral venous blood samples were collected at three time intervals post symptom onset (PSO): 1-3 mo, 4-8 mo, and 9-12 mo. The humoral immune response was evaluated by measuring anti-SARS-CoV-2 IgG, virus-neutralizing antibody activity, total S1-specific B-cells, and B cell subpopulations. Results: The levels of anti-SARS-CoV-2 specific IgG antibodies decreased from 390.3 to 204.5 BAU/ml in the first 6-8 months PSO but did not significantly decrease further until the 12 th mo (126.6 BAU/ml). Virus-neutralizing antibodies (activity decreased by 20.4% between the 1st and 6-8th mos but remained relatively stable thereafter and could be detected up to 12 months PSO. In peripheral blood, the amount of S1-specific plasmablasts was highest one month after COVID-19 infection, and the level of memory B cells at 6 months. Those were detected even 12 months PSO, albeit in smaller quantities.  Conclusion: The study provides evidence for the persistence of SARS-CoV-2-specific B-cell immune memory up to 1year post-infection. The presence of virus-specific memory B cells and plasmablasts suggests potential for sustained protection against reinfection. Further research is needed to elucidate the role of B-cell immune memory in preventing infection and to understand the individual variations of immune response

A comparative study between children and adults with bacterial neuroinfections

Abstract Introduction: Bacterial meningitis is an acute purulent infection of the meninges. There are significant differences in the etiological spectrum, clinical course and outcome of bacterial meningitis in the age groups, and their recognition is important for early diagnosis and adequate therapy. Aim: The study aims to determine the differences in the etiology and clinical presentation of bacterial meningitis between children and adults. Materials and methods: The study included 90 patients (25 children and 65 adults) with bacterial neuroinfection admitted to St George University Hospital, Plovdiv between January 1, 2016 and September 31, 2019. We applied epidemiological and clinical analysis, microbiological and statistical methods. Results: In adults, the most common etiological agent was Streptococcus pneumoniae (20%), followed by Staphylococcus spp. (18.5%), Listeria monocytogenes (12.3%), Streptococcus spp. (3.1%), Haemophilus influenzae (3.1%), Klebsiella pneumoniae (1.5%), and Mycobacterium tuberculosis (1.5%). The etiological structure in children was different: Neisseria meningitidis (20%), Streptococcus pneumoniae (16%), Klebsiella pneumoniae (8%), Enterococcus faecium (8%), Streptococcus salivarius (4%), and Mycobacterium tuberculosis (4%). In 40% of the cases, both children and adults, the causative agent was not identified. Conclusions: Regarding the clinical presentation, a statistical significance between the age groups was found with headache and alterations in consciousness, more commonly seen in adults, while vomiting, ear pain was more common in children (p<0.05). Concomitant otitis, sinusitis, pneumonia, and sepsis were often observed. The mortality rate was much higher in adults (43%) when compared with children (8%)

Cannabidiol improves memory and decreases IL-1β serum levels in rats with lipopolysaccharide-induced inflammation

Aim: Memory improving and anti-inflammatory properties of cannabidiol (CBD) were investigated in an experimental model of lipopolysaccharide (LPS)-induced inflammation. Materials and methods: Male Wistar rats were randomly divided into 4 groups: control, LPS control, LPS + CBD 5 mg/kg bw, and LPS + CBD 10 mg/kg bw. Animals were treated with CBD 14 days before LPS administration and throughout the experiment. Step-through passive avoidance task, Y-maze, and novel object recognition test (NORT) were used to assess the memory functions. The following parameters were recorded: latency time, spontaneous alternations percentage (SA%) and recognition index (RI). IL-10, IL-6, TNF-α, and IL-1β serum levels were measured to evaluate the immunomodulatory properties of CBD. Results: LPS led to significant decrease of the recorded parameters in all memory tasks. This demonstrated the memory-impairing effect of LPS-induced inflammation. In the Y-maze and NORT tests, both doses of CBD increased SA% and RI, respectively. Significant difference was found in comparison with the LPS controls. Rats from the CBD treated groups showed increased latency in the step-through passive avoidance task. In the short-term memory test, both CBD doses significantly increased this parameter when compared with both control groups (p<0.05 and p<0.001, respectively), whereas in the long-term memory test, statistical significance was reached only in comparison with the LPS controls (p<0.01). CBD treatment failed to reduce TNF-α and IL-6 serum levels. The lower studied dose significantly decreased IL-10 and IL-1β concentrations compared to LPS controls (p<0.01 and p<0.05, respectively). Conclusions: CBD improved spatial working and recognition memory in rats with LPS-induced inflammation. Suppression of IL-1β production could be attributed to the observed effect

Potential value of a rapid syndromic multiplex PCR for the diagnosis of native and prosthetic joint infections: a real-world evidence study

Introduction: The BIOFIRE Joint Infection (JI) Panel is a diagnostic tool that uses multiplex-PCR testing to detect microorganisms in synovial fluid specimens from patients suspected of having septic arthritis (SA) on native joints or prosthetic joint infections (PJIs). Methods: A study was conducted across 34 clinical sites in 19 European and Middle Eastern countries from March 2021 to June 2022 to assess the effectiveness of the BIOFIRE JI Panel. Results: A total of 1527 samples were collected from patients suspected of SA or PJI, with an overall agreement of 88.4 % and 85 % respectively between the JI Panel and synovial fluid cultures (SFCs). The JI Panel detected more positive samples and microorganisms than SFC, with a notable difference on Staphylococcus aureus, Streptococcus species, Enterococcus faecalis, Kingella kingae, Neisseria gonorrhoeae, and anaerobic bacteria. The study found that the BIOFIRE JI Panel has a high utility in the real-world clinical setting for suspected SA and PJI, providing diagnostic results in approximately 1 h. The user experience was positive, implying a potential benefit of rapidity of results' turnover in optimising patient management strategies. Conclusion: The study suggests that the BIOFIRE JI Panel could potentially optimise patient management and antimicrobial therapy, thus highlighting its importance in the clinical setting

Care of patients with inborn errors of immunity in thirty J Project countries between 2004 and 2021

IntroductionThe J Project (JP) physician education and clinical research collaboration program was started in 2004 and includes by now 32 countries mostly in Eastern and Central Europe (ECE). Until the end of 2021, 344 inborn errors of immunity (IEI)-focused meetings were organized by the JP to raise awareness and facilitate the diagnosis and treatment of patients with IEI.ResultsIn this study, meeting profiles and major diagnostic and treatment parameters were studied. JP center leaders reported patients’ data from 30 countries representing a total population of 506 567 565. Two countries reported patients from JP centers (Konya, Turkey and Cairo University, Egypt). Diagnostic criteria were based on the 2020 update of classification by the IUIS Expert Committee on IEI. The number of JP meetings increased from 6 per year in 2004 and 2005 to 44 and 63 in 2020 and 2021, respectively. The cumulative number of meetings per country varied from 1 to 59 in various countries reflecting partly but not entirely the population of the respective countries. Altogether, 24,879 patients were reported giving an average prevalence of 4.9. Most of the patients had predominantly antibody deficiency (46,32%) followed by patients with combined immunodeficiencies (14.3%). The percentages of patients with bone marrow failure and phenocopies of IEI were less than 1 each. The number of patients was remarkably higher that those reported to the ESID Registry in 13 countries. Immunoglobulin (IgG) substitution was provided to 7,572 patients (5,693 intravenously) and 1,480 patients received hematopoietic stem cell therapy (HSCT). Searching for basic diagnostic parameters revealed the availability of immunochemistry and flow cytometry in 27 and 28 countries, respectively, and targeted gene sequencing and new generation sequencing was available in 21 and 18 countries. The number of IEI centers and experts in the field were 260 and 690, respectively. We found high correlation between the number of IEI centers and patients treated with intravenous IgG (IVIG) (correlation coefficient, cc, 0,916) and with those who were treated with HSCT (cc, 0,905). Similar correlation was found when the number of experts was compared with those treated with HSCT. However, the number of patients treated with subcutaneous Ig (SCIG) only slightly correlated with the number of experts (cc, 0,489) and no correlation was found between the number of centers and patients on SCIG (cc, 0,174).Conclusions1) this is the first study describing major diagnostic and treatment parameters of IEI care in countries of the JP; 2) the data suggest that the JP had tremendous impact on the development of IEI care in ECE; 3) our data help to define major future targets of JP activity in various countries; 4) we suggest that the number of IEI centers and IEI experts closely correlate to the most important treatment parameters; 5) we propose that specialist education among medical professionals plays pivotal role in increasing levels of diagnostics and adequate care of this vulnerable and still highly neglected patient population; 6) this study also provides the basis for further analysis of more specific aspects of IEI care including genetic diagnostics, disease specific prevalence, newborn screening and professional collaboration in JP countries