Quantification of the HIV-1 total reservoir in the peripheral blood of naïve and treated patients by a standardised method derived from a commercial HIV-1 RNA quantification assay.

OBJECTIVES: HIV-1 DNA can persist in host cells, establishing a latent reservoir. This study was aimed to develop an extraction and amplification protocol for HIV-1 DNA quantification by modifying a quantitative commercial assay. METHODS: HIV-1 DNA was extracted on an Abbott m2000sp instrument, using an open-mode protocol. Two calibrators, spiked with a plasmid containing HIV-1 genome (103 and 105 cps/mL), were extracted and amplified to generate a master calibration curve. Precision, accuracy, linear dynamic range, limit of quantification (LOQ) and limit of detection (LOD) were determined. A cohort of patients, naïve or chronically infected, was analysed. RESULTS: Calibration curve was obtained from 42 replicates of standards (stds); precision was calculated (coefficients of variability [CVs] below 10%); accuracy was higher than 90%. Linearity covered the entire range tested (10-104 copies per reaction), and LOD (95%) was 12 copies per reaction. HIV-1 DNA was significantly higher (p < 0.0001) in drug-naïve (62) than in chronically treated patients (50), and proviral loads correlated with lymphocytes (p = 0.0002) and CD4+ (p < 0.0001) counts only in naïve patients. Both groups displayed a significant inverse correlation between CD4+ nadir and proviral loads. A significant correlation (p < 0.0001) between viraemia and HIV-1 reservoir was disclosed. No significant difference was obtained from the comparison between proviral loads on whole blood and peripheral blood mononuclear cells (PBMCs) from the same patient. CONCLUSIONS: The novelty of our approach relies on the selection of appropriate reference standard extracted and amplified as clinical specimens avoiding any underestimation of the reservoir. Results confirm HIV-1 DNA as a marker of disease progression, supporting the relationship between the width of latent reservoir and the immunological status of the patient.

Fatal anaphylaxis in Italy: Analysis of cause-of-death national data, 2004-2016.

BACKGROUND: Epidemiological data on fatal anaphylaxis are underestimated worldwide. Few Italian data do exist. The aims of the study are to determine the anaphylaxis mortality rate in Italy and its associations with demographic characteristics (gender, age, and geographical distribution), and to investigate which are the most common triggers of fatal anaphylaxis. MATERIAL AND METHODS: This is a descriptive study analyzing data reported to the National Register of Causes of Death database and managed by the Italian National Institute of Statistics for the years 2004-2016. An analytical method was developed to identify all the ICD-10 codes related to anaphylaxis deaths, which were divided into two classes: "Definite anaphylaxis deaths" and "Possible anaphylaxis deaths." RESULTS: From 2004 through 2016, 392 definite anaphylaxis deaths and 220 possible anaphylaxis deaths were recorded. The average mortality rate for definite anaphylaxis, from 2004 to 2016, was 0.51 per million population per year. Definite fatal anaphylaxis was mostly due to the use of medications (73.7%), followed by unspecified causes (20.7%) and hymenoptera stings (5.6%). Concerning possible anaphylaxis deaths, the most common cause was venom-stinging insect (51.4%). We did not find any data on food fatal anaphylaxis. Unspecified anaphylaxis accounted for 21%-28% of all cases, underlining the difficulty in accurately ascertaining the causes of fatal anaphylaxis and therefore in assigning the proper ICD-10 code. CONCLUSION: This is the first study of anaphylaxis-related mortality coming from an official database of the whole Italian population. However, the actual number of deaths by anaphylaxis, and their related triggers, is probably underreported, mostly due to limitations of the current recording system, and to a poor allergy education. Corrective actions should be undertaken for the benefit of the Health System.

Development of a model care pathway for the management of Hymenoptera venom allergy: evidence-based key interventions and indicators.

BACKGROUND: Hymenoptera venom allergy (HVA) is an underestimated condition representing an important cause of morbidity and mortality worldwide. Preventing future allergic reactions in patients who have already developed a systemic reaction is based on the correct management of the acute phase of the reaction followed by a correct diagnosis and, where indicated, prescription of adrenaline autoinjectors and VIT. A possible strategy to optimize care processes and to improve outcomes is the implementation of a Diagnostic and Therapeutic Care Pathways, also known as Integrated Care Pathways or Clinical Pathways (CPWs). The aim of the care pathway is to enhance the quality of care by improving risk-adjusted patient outcomes, promoting patient safety, increasing patient satisfaction, and optimizing the use of resources. To our knowledge, currently in Italy as well as in Europe, there is no CPWs codified for the management of HVA patients. This paper describes the development of the clinical content of a care pathway for the management of HVA. METHODS: The methodology applied is based on the eight step method to build the clinical content of an evidence-based care pathway suggested by Lodewijckx et al. RESULTS: Three hundred and seventeen different clinical activities were extracted from the selected literature. The expert panel was involved in their evaluation, expressing a judgment of relevance through the Delphi study. As a result, 126 clinical activities were appraised to be valid and feasible. The final recommendations (126) were translated into 123 key interventions. Six indicators were produced by the clinical activities. CONCLUSION: A set of 123 key interventions and of six process indicators were found to be appropriate for the development and standardization of the clinical content of the Hymenoptera venom allergy care pathway.