A retrospective observational study of 1000 consecutive patients tested with the FilmArray® Meningitis/Encephalitis panel: clinical diagnosis at discharge and microbiological findings.

FilmArray® Meningitis/Encephalitis panel (FAME-p) is used to diagnose central nervous system (CNS) infections. In this study, we investigated performance of FAME-p compared to comparator assays (CA), and for the first time, clinical diagnosis at discharge (CDD). 1000 consecutive patients with a cerebrospinal fluid (CSF) sample analyzed with FAME-p were identified. As CA, culture, polymerase chain reaction and cryptococcal antigen test were used. Medical records of patients were obtained. A CDD of CNS infection was made in 139 of 1000 CSF samples. FAME-p was positive in 66 samples with 44 viral and 22 bacterial agents. Thirteen FAME-p findings were not confirmed by CA, with four discrepant results remaining after comparison with the CDD. Positive percentage agreement (PPA) calculated against CA was 100%. Negative percentage agreement (NPA) calculated against CA was 94.4-99.8% for Haemophilus influenzae, Listeria monocytogenes, Streptococcus agalactiae, S. pneumoniae and varicella-zoster virus (VZV). NPA calculated against CDD was higher (compared to CA) for L. monocytogenes, S. agalactiae and VZV (100%), and lower for Escherichia coli, enterovirus and herpes simplex virus 2 (50-83.3%). NPA of FAME-p for human herpes virus 6 was difficult to interpret. Eighty-four cases received diagnosis of CNS-infection despite negative FAME-p. The four most common non-infectious etiologies were primary headache disorders, cranial nerve palsies, neuroinflammatory disorders and seizure. Although FAME-p shows good performance in diagnosis of CNS infections, result of FAME-p should be interpreted carefully. Considering infectious diseases not covered by FAME-p as well as non-infectious differential diagnoses is important in this context.

A case report of a chronic migraine patient treated with three different anti-CGRP monoclonal antibodies: which parameters better represent the efficacy?

Objective: To report the efficacy of different anti-calcitonin gene-related peptide (CGRP) monoclonal antibodies (mAbs) on headache frequency, intensity, and duration. Background: Blockade of CGRP receptors or neuropeptide with anti-CGRP mAbs have been successfully used for several years for the prevention of chronic and episodic migraine. The response is usually assessed by improvement seen in the number of days with headache per month. However, clinical praxis indicates that sole reliance on the frequency of headaches might be insufficient to interpret the efficacy of these treatments. Methods: Retrospective review of a case with a meticulous headache diary who has tried three different anti-CGRP mAbs for chronic migraine prevention. Results: The patient has been diagnosed with chronic migraine and was first treated with erenumab, followed by fremanezumab and thereafter galcanezumab due to several reasons. In addition to significant improvement in all three parameters analyzed with anti-CGRP mAb treatment, the most important and valuable effect on the patient's quality of life was decreased duration and frequency of headaches. At present, the patient is receiving fremanezumab treatment with an excellent tolerability. Conclusion: There is a clear need for careful follow-up and detailed daily records of headaches showing the frequency, duration, and severity for the evaluation of anti-CGRP mAbs treatment. This study shows the importance of this information in order for medical professionals to make an informed decision regarding the best course of anti-CGRP mAbs treatment in cases of side effects or lack of efficacy.

Trends in Antimicrobial Drug Resistance of Streptococcus pneumoniae Isolates at Jordan University Hospital (2000⁻2018).

Antimicrobial drug resistance (AMR) in pneumococci complicates the treatment of serious pneumococcal infections. Country-specific AMR patterns can help to establish guidelines for empiric therapy. The aim of the current study was to analyze the distribution of AMR among Streptococcus pneumoniae isolates at Jordan University Hospital (JUH) during 2000⁻2018. Paper-based and electronic clinical data registry records from 2000 to 2018 were retrospectively analyzed to study the AMR among pneumococcal isolates at JUH. Temporal trend analysis was done using two-tailed linear-by-linear test for association. The total number of unique pneumococcal isolates that were identified was 556, of which 544 isolates had antimicrobial susceptibility testing results. The most frequent specimens were eye (n = 117, 21.0%), bloodstream (n = 93, 16.7%) and sputum (n = 81, 14.6%). Invasive infections represented 23.6% of all unique isolates. The overall susceptibility of S. pneumoniae isolates during the study period to different antimicrobials was: 100% to vancomycin, 97.7% to ceftriaxone, 97.1% to cefotaxime, 94.9% to chloramphenicol, 89.7% to penicillin, 83.8% to levofloxacin, 67.7% to clindamycin and 52.1% to erythromycin. The prevalence of multi-drug resistance (MDR) was 8.6% (95% confidence interval: 6.4⁻11.5%). Trend analysis showed an increase in the prevalence of non-susceptibility to erythromycin, clindamycin and levofloxacin (p < 0.001). MDR prevalence increased from 1.6% in the first quarter to 14.6% in the fourth quarter (p < 0.001). The incidence of invasive infections declined over the study period (p < 0.001). The increase in the prevalence of AMR and MDR among pneumococcal isolates in Jordan demands judicious use of antimicrobials and regular surveillance of resistance.

Automated image-based assay for evaluation of HIV neutralization and cell-to-cell fusion inhibition.

BACKGROUND: Standardized techniques to detect HIV-neutralizing antibody responses are of great importance in the search for an HIV vaccine. METHODS: Here, we present a high-throughput, high-content automated plaque reduction (APR) assay based on automated microscopy and image analysis that allows evaluation of neutralization and inhibition of cell-cell fusion within the same assay. Neutralization of virus particles is measured as a reduction in the number of fluorescent plaques, and inhibition of cell-cell fusion as a reduction in plaque area. RESULTS: We found neutralization strength to be a significant factor in the ability of virus to form syncytia. Further, we introduce the inhibitory concentration of plaque area reduction (ICpar) as an additional measure of antiviral activity, i.e. fusion inhibition. CONCLUSIONS: We present an automated image based high-throughput, high-content HIV plaque reduction assay. This allows, for the first time, simultaneous evaluation of neutralization and inhibition of cell-cell fusion within the same assay, by quantifying the reduction in number of plaques and mean plaque area, respectively. Inhibition of cell-to-cell fusion requires higher quantities of inhibitory reagent than inhibition of virus neutralization.

Frequent intratype neutralization by plasma immunoglobulin a identified in HIV type 2 infection.

Human immunodeficiency virus type 2 (HIV-2) is less transmissible and less pathogenic compared to HIV-1 and, when matched for CD4(+) T cell count, the plasma viral load in HIV-2-infected individuals is approximately one log lower than in HIV-1-infected individuals. The explanation for these observations is elusive, but differences in virus controlling immunity generated in the two infections may be contributing factors. In the present study, we investigated neutralization by immunoglobulin A (IgA), in parallel with IgG, purified from plasma of HIV-1, HIV-2, and HIV-1/HIV-2 dually (HIV-D) infected individuals. Neutralization was analyzed against HIV-1 and HIV-2 isolates using a plaque reduction assay. In HIV-2 infection, intratype-specific neutralization by IgA was frequently detected, although at a lesser magnitude then the corresponding IgG neutralizing titers. In contrast, neutralization by IgA could rarely be demonstrated in HIV-1 infection despite similar plasma IgA levels in both infections. In addition, IgA and IgG of HIV-D plasma neutralized the HIV-2 isolate more potently than the HIV-1 isolate, suggesting that the difference between neutralizing activity of plasma IgA and IgG depends on the virus itself. Taken together, these findings suggest that both IgA and IgG add to the potent intratype neutralizing activity detected in HIV-2 plasma, which may contribute to virus control in HIV-2 infection.

Effect of complement on HIV-2 plasma antiviral activity is intratype specific and potent.

Human immunodeficiency virus type 2 (HIV-2)-infected individuals develop immunodeficiency with a considerable delay and transmit the virus at rates lower than HIV-1-infected persons. Conceivably, comparative studies on the immune responsiveness of HIV-1- and HIV-2-infected hosts may help to explain the differences in pathogenesis and transmission between the two types of infection. Previous studies have shown that the neutralizing antibody response is more potent and broader in HIV-2 than in HIV-1 infection. In the present study, we have examined further the function of the humoral immune response and studied the effect of complement on the antiviral activity of plasma from singly HIV-1- or HIV-2-infected individuals, as well as HIV-1/HIV-2 dually infected individuals. The neutralization and antibody-dependent complement-mediated inactivation of HIV-1 and HIV-2 isolates were tested in a plaque reduction assay using U87.CD4.CCR5 cells. The results showed that the addition of complement increased intratype antiviral activities of both HIV-1 and HIV-2 plasma samples, although the complement effect was more pronounced with HIV-2 than HIV-1 plasma. Using an area-under-the-curve (AUC)-based readout, multivariate statistical analysis confirmed that the type of HIV infection was independently associated with the magnitude of the complement effect. The analyses carried out with purified IgG indicated that the complement effect was largely exerted through the classical complement pathway involving IgG in both HIV-1 and HIV-2 infections. In summary, these findings suggest that antibody binding to HIV-2 structures facilitates the efficient use of complement and thereby may be one factor contributing to a strong antiviral activity present in HIV-2 infection.

Potent intratype neutralizing activity distinguishes human immunodeficiency virus type 2 (HIV-2) from HIV-1.

HIV-2 has a lower pathogenicity and transmission rate than HIV-1. Neutralizing antibodies could be contributing to these observations. Here we explored side by side the potency and breadth of intratype and intertype neutralizing activity (NAc) in plasma of 20 HIV-1-, 20 HIV-2-, and 11 dually HIV-1/2 (HIV-D)-seropositive individuals from Guinea-Bissau, West Africa. Panels of primary isolates, five HIV-1 and five HIV-2 isolates, were tested in a plaque reduction assay using U87.CD4-CCR5 cells as targets. Intratype NAc in HIV-2 plasma was found to be considerably more potent and also broader than intratype NAc in HIV-1 plasma. This indicates that HIV-2-infected individuals display potent type-specific neutralizing antibodies, whereas such strong type-specific antibodies are absent in HIV-1 infection. Furthermore, the potency of intratype NAc was positively associated with the viral load of HIV-1 but not HIV-2, suggesting that NAc in HIV-1 infection is more antigen stimulation dependent than in HIV-2 infection, where plasma viral loads typically are at least 10-fold lower than in HIV-1 infection. Intertype NAc of both HIV-1 and HIV-2 infections was, instead, of low potency. HIV-D subjects had NAc to HIV-2 with similar high potency as singly HIV-2-infected individuals, whereas neutralization of HIV-1 remained poor, indicating that the difference in NAc between HIV-1 and HIV-2 infections depends on the virus itself. We suggest that immunogenicity and/or antigenicity, meaning the neutralization phenotype, of HIV-2 is distinct from that of HIV-1 and that HIV-2 may display structures that favor triggering of potent neutralizing antibody responses.

Generation of neutralizing antibodies and divergence of SIVmac239 in cynomolgus macaques following short-term early antiretroviral therapy.

Neutralizing antibodies (NAb) able to react to heterologous viruses are generated during natural HIV-1 infection in some individuals. Further knowledge is required in order to understand the factors contributing to induction of cross-reactive NAb responses. Here a well-established model of experimental pathogenic infection in cynomolgus macaques, which reproduces long-lasting HIV-1 infection, was used to study the NAb response as well as the viral evolution of the highly neutralization-resistant SIVmac239. Twelve animals were infected intravenously with SIVmac239. Antiretroviral therapy (ART) was initiated ten days post-inoculation and administered daily for four months. Viral load, CD4(+) T-cell counts, total IgG levels, and breadth as well as strength of NAb in plasma were compared simultaneously over 14 months. In addition, envs from plasma samples were sequenced at three time points in all animals in order to assess viral evolution. We report here that seven of the 12 animals controlled viremia to below 10(4) copies/ml of plasma after discontinuation of ART and that this control was associated with a low level of evolutionary divergence. Macaques that controlled viral load developed broader NAb responses early on. Furthermore, escape mutations, such as V67M and R751G, were identified in virus sequenced from all animals with uncontrolled viremia. Bayesian estimation of ancestral population genetic diversity (PGD) showed an increase in this value in non-controlling or transient-controlling animals during the first 5.5 months of infection, in contrast to virus-controlling animals. Similarly, non- or transient controllers displayed more positively-selected amino-acid substitutions. An early increase in PGD, resulting in the generation of positively-selected amino-acid substitutions, greater divergence and relative high viral load after ART withdrawal, may have contributed to the generation of potent NAb in several animals after SIVmac239 infection. However, early broad NAb responses correlated with relatively preserved CD4(+) T-cell numbers, low viral load and limited viral divergence.