Elucidating the 3D Structure of a Surface Membrane Antigen from Trypanosoma cruzi as a Serodiagnostic Biomarker of Chagas Disease.

Chagas disease (CD) is a vector-borne parasitosis, caused by the protozoan parasite Trypanosoma cruzi, that affects millions of people worldwide. Although endemic in South America, CD is emerging throughout the world due to climate change and increased immigratory flux of infected people to non-endemic regions. Containing of the diffusion of CD is challenged by the asymptomatic nature of the disease in early infection stages and by the lack of a rapid and effective diagnostic test. With the aim of designing new serodiagnostic molecules to be implemented in a microarray-based diagnostic set-up for early screening of CD, herein, we report the recombinant production of the extracellular domain of a surface membrane antigen from T. cruzi (TcSMP) and confirm its ability to detect plasma antibodies from infected patients. Moreover, we describe its high-resolution (1.62 Å) crystal structure, to which in silico epitope predictions were applied in order to locate the most immunoreactive regions of TcSMP in order to guide the design of epitopes that may be used as an alternative to the full-length antigen for CD diagnosis. Two putative, linear epitopes, belonging to the same immunogenic region, were synthesized as free peptides, and their immunological properties were tested in vitro. Although both peptides were shown to adopt a structural conformation that allowed their recognition by polyclonal antibodies raised against the recombinant protein, they were not serodiagnostic for T. cruzi infections. Nevertheless, they represent good starting points for further iterative structure-based (re)design cycles.

Structure, Immunoreactivity, and In Silico Epitope Determination of SmSPI S. mansoni Serpin for Immunodiagnostic Application.

The human parasitic disease Schistosomiasis is caused by the Schistosoma trematode flatworm that infects freshwaters in tropical regions of the world, particularly in Sub-Saharan Africa, South America, and the Far-East. It has also been observed as an emerging disease in Europe, due to increased immigration. In addition to improved therapeutic strategies, it is imperative to develop novel, rapid, and sensitive diagnostic tests that can detect the Schistosoma parasite, allowing timely treatment. Present diagnosis is difficult and involves microscopy-based detection of Schistosoma eggs in the feces. In this context, we present the 3.22 Å resolution crystal structure of the circulating antigen Serine protease inhibitor from S. mansoni (SmSPI), and we describe it as a potential serodiagnostic marker. Moreover, we identify three potential immunoreactive epitopes using in silico-based epitope mapping methods. Here, we confirm effective immune sera reactivity of the recombinant antigen, suggesting the further investigation of the protein and/or its predicted epitopes as serodiagnostic Schistosomiasis biomarkers.

Case Report: A Fatal Case of West Nile Virus Meningoencephalomyelitis in a Woman with Systemic Lupus Erythematosus Initially Misdiagnosed as SARS-CoV-2 Infection.

We present a fatal case of West Nile virus meningoencephalomyelitis initially misdiagnosed as COVID-19 in a 63-year-old Egyptian woman with a previous diagnosis of systemic lupus erythematosus. The patient's medical history and immunosuppressive therapy, as well as the COVID-19 pandemic, substantially broadened the differential diagnosis of her encephalitis.

Imported dengue fever: a 16-years retrospective analysis in Milan (Italy) and a brief review of the European literature.

Dengue Fever (DF), transmitted by Aedes mosquitoes, is the most common arthropod-borne infection, it is almost ubiquitous in tropical and subtropical areas with an estimate of 360 million infections per year. A competent vector (A. albopictus) is present in most of Southern Europe and is endemic in Italy. We conducted a 16-year retrospective study of probable/confirmed dengue fever observed at the Department of Infectious Diseases of Luigi Sacco Hospital in Milan, Italy. Overall 122 patients were included in the study, 106 with probable and 16 with proven diagnosis of dengue fever. Most patients (91%) were Italian, with a median age of 35 years (IQR 29-46 years) and similar gender distribution, travelling for tourism (80%). Asia (mainly South East Asia and Indian Subcontinent) was the most frequent travel destination (55%), followed by Central America and the Caribbeans (22%). August-September was the peak season of presentation (42.6%). The majority of our diagnoses were based on serology alone. The most common signs and symptoms were fever (99,2%), maculopapular rash (50,8%), headache (50,8%), arthralgias (50,8%) and myalgias (46,7%). Leukopenia (77%), thrombocytopenia (81%) and altered LDH, AST and ALT (respectively 60,6%, 54,1% and 45,9%) were the most common laboratory test's abnormalities. No cases of severe DF were recorded. Our epidemiological and clinical findings are largely in accordance with most recent studies about imported DF in Europe. Although very similar in presentation to other arthropod-borne illnesses, some clinical features may help in differentiating DF from other causes of fever in the returning traveler.

Dengue fever complicated by liver dysfunction due to possible co-infection with hepatitis E in a returning traveller from Cuba.

Dengue fever is a mosquito-borne infection that co-circulates with Chikungunya and Zika virus infection in many parts of the world. Dengue virus (DENV) is occasionally responsible for acute hepatitis and a few cases of acute hepatitis due to co-infection with DENV and hepatitis E virus have been described in India. A 37-year-old Cuban woman living in Italy was admitted to our hospital with a presumed arboviral infection upon her return to Italy short after a 15-day trip to her home-country to visit relatives. An acute infection due to DENV serotype 1 was initially diagnosed, following a clinical course characterized by signs of liver dysfunction that were possibly due to co-infection with hepatitis E virus.

Primary HIV infection in patients with acute hepatitis B: a report of two cases.

We describe two patients admitted to our institution with a diagnosis of sexually acquired acute hepatitis B who also had underlying hyper acute HIV infection. Both individuals reported high rates of condomless sex. Antiviral therapy active against HBV and HIV was started within days after diagnosis. Treatment was well tolerated and led to a rapid control of both infections and hepatitis B surface antibody seroconversion. The efficacy and safety of contemporary antiretroviral drug combinations suggest that treatment of acute HIV infection is feasible in patients with acute hepatitis B.

HIV avidity index performance using a modified fourth-generation immunoassay to detect recent HIV infections.

BACKGROUND: Detecting recent HIV infections is important to evaluate incidence and monitor epidemic trends. We aimed to evaluate the diagnostic performance and accuracy of the avidity index (AI) for discriminating for recent HIV infections. METHODS: We collected serum samples from HIV-1 positive individuals: A) with known date of infection (midpoint in time between last HIV-negative and first HIV-positive test); B) infected for >1 year. Samples were divided into two aliquots: one diluted with phosphate buffered saline (PBS) and the other with 1 M guanidine. Both aliquots were assayed by the Architect HIV Ag/Ab Combo 4th generation assay (Abbott). We compared AI found in recent (RI=<6 months from seroconversion) and established (EI) infections. The diagnostic accuracy was evaluated by receiver operating characteristic (ROC) curve analysis. The proportion of samples misclassified as recent (FRR) was calculated. RESULTS: In total, 647 samples were collected: 455 in group A (51.6% RI and 48.4% EI) and 192 in group B. Among these, sixteen samples were from elite controllers, 294 from treated patients, 328 from patients infected with non-B subtypes. Samples before antiretroviral initiation showed a mean AI significantly lower among RI compared to EI (0.66+0.28 vs. 1.00±0.12; p<0.000). The FRR was 0% using a cut-off of ≤0.70. An extremely low FRR was observed among elite controllers, samples with low VL or CD4. HIV subtype had no impact on AI misclassifications. All individuals in group A reached the AI threshold of 0.80 within 24 months after seroconversion. CONCLUSIONS: The AI is an accurate serological marker for discriminating recent from established HIV infections and meets WHO requirements for HIV incidence assays.

Imported arboviral infections in Italy, July 2014-October 2015: a National Reference Laboratory report.

BACKGROUND: Imported cases of infections due to Dengue (DENV) and Chikungunya (CHIKV) viruses and, more recently, Zika virus (ZIKV) are commonly reported among travelers returning from endemic regions. In areas where potentially competent vectors are present, the risk of autochthonous transmission of these vector-borne pathogens is relatively high. Laboratory surveillance is crucial to rapidly detect imported cases in order to reduce the risk of transmission. This study describes the laboratory activity performed by the National Reference Laboratory for Arboviruses (NRLA) at the Italian National Institute of Health in the period from July 2014 to October 2015. METHODS: Samples from 180 patients visited/hospitalized with a suspected DENV/CHIKV/ZIKV infection were sent to the NRLA from several Italian Hospitals and from Regional Reference Laboratories for Arboviruses, in agreement with the National Plan on human surveillance of vector-borne diseases. Both serological (ELISA IgM test and Plaque Reduction Neutralization Test-PRNT) and molecular assays (Real Time PCR tests, RT-PCR plus nested PCR and sequencing of positive samples) were performed. RESULTS: DENV infection was the most frequently diagnosed (80 confirmed/probable cases), and all four genotypes were detected. However, an increase in imported CHIKV cases (41 confirmed/probable cases) was observed, along with the detection of the first ZIKV cases (4 confirmed cases), as a consequence of the recent spread of both CHIKV and ZIKV in the Americas. CONCLUSIONS: Main diagnostic issues highlighted in our study are sensitivity limitations of molecular tests, and the importance of PRNT to confirm serological results for differential diagnosis of Arboviruses. The continuous evaluation of diagnostic strategy, and the implementation of laboratories networks involved in surveillance activities is essential to ensure correct diagnosis, and to improve the preparedness for a rapid and proper identification of viral threats.

Pregnancy-related changes of antiretroviral pharmacokinetics: an argument for therapeutic drug monitoring.

Here we describe a case of an HIV-infected young woman with extensive drug-resistant virus, who was successfully switched from a raltegravir-based regimen to a dolutegravir-based intensified antiretroviral regimen a few days before scheduled caesarean section because of the still detectable viral load. The trough concentrations of all antiretroviral drugs before and after delivery are also described. Our case underlines both the difficult management of young women, HIV-infected at young age with very limited treatment options and the great variability in the pregnancy-related physiological changes affecting the pharmacokinetics of antiretrovirals.

Unexpected vertical transmission of HIV infection.

Mother-to-child transmission of HIV infection occurred in a child born from an HIV-infected mother with HIV-RNA undetectable during pregnancy. She was suffering from gastroenteritis in the last 3 weeks of gestation.

Epidemiological and clinical characteristics of pediatric gastroenteritis associated with new viral agents.

A 22-month study (2008-2009) was carried out on 273 patients (average age 40 months), admitted with gastroenteritis to the Pediatric Unit of L. Sacco University Hospital in Milan, Italy. Fecal samples were investigated for rotavirus (HRV), norovirus (NoV), adenovirus (AdV), sapovirus (SaV), enterovirus, astrovirus and bocavirus (HBoV). A 38.3% incidence of infection was observed for HRV, followed by NoV (16.2%), HBoV (13.6%), AdV (2.6%) and SaV (0.6%). Clinical evaluation of 109 gastroenteritis patients with confirmed diagnosis was graded by the Ruska-Vesikari scoring system, showing vomiting (78%), diarrhea (96%) and fever (80%). A total of 25 NoV-positive samples were selected for nucleotide sequence analysis. The severity of AdV-associated infection was lower than for NoV, HRV and HBoV. These latter viruses caused similar symptoms that were indistinguishable using clinical information. NoV, HRV and HBoV were often present as mixed infections (13.1%). Sequencing of NoV-positive samples allowed identification of GII.2, GII.3 and GII.4 2006 variants.