Severe West Nile Virus Neuroinvasive Disease: Clinical Characteristics, Short- and Long-Term Outcomes.

West Nile Virus Neuroinvasive Disease (WNV NID) requires prolonged intensive care treatment, resulting in high mortality and early disability. Long-term results are lacking. We have conducted an observational retrospective study with a prospective follow-up of WNV NID patients treated at the Intensive Care Unit (ICU), University Hospital for Infectious Diseases, Zagreb, Croatia, 2013-2018. Short-term outcomes were vital status, length of stay (LOS), modified Rankin Scale (mRS), and disposition at discharge. Long-term outcomes were vital status and mRS at follow-up. Twenty-three patients were identified, 78.3% males, median age 72 (range 33-84) years. Two patients (8.7%) died in the ICU, with no lethal outcomes after ICU discharge. The median ICU LOS was 19 days (range 5-73), and the median hospital LOS was 34 days (range 7-97). At discharge, 15 (65.2%) patients had moderate to severe/mRS 3-5, 6 (26.0%) had slight disability/mRS 2-1, no patients were symptom-free/mRS 0. Ten (47.6%) survivors were discharged to rehabilitation facilities. The median time to follow-up was nine months (range 6-69). At follow-up, seven patients died (30.5%), five (21.7%) had moderate to severe/mRS 3-5, one (4.3%) had slight disability/mRS 2-1, six (26.1%) had no symptoms/mRS 0, and four (17.4%) were lost to follow-up. Briefly, ten (43.5%) survivors improved their functional status, one (4.3%) was unaltered, and one (4.3%) aggravated. In patients with severe WNV NID, intensive treatment in the acute phase followed by inpatient rehabilitation resulted in significant recovery of functional status after several months.

Neutralizing Antibodies against SARS-CoV-2 Beta and Omicron Variants Inhibition Comparison after BNT162b2 mRNA Booster Doses with a New PETIA sVNT Assay

Background: Monitoring antibody response following SARS-CoV-2 vaccination is strategic, and neutralizing antibodies represent the gold standard. The neutralizing response to Beta and Omicron VOCs was evaluated versus the gold standard by a new commercial automated assay. Methods: Serum samples from 100 healthcare workers from the Fondazione Policlinico Universitario Campus Biomedico and the Pescara Hospital were collected. IgG levels were determined by chemiluminescent immunoassay (Abbott Laboratories, Wiesbaden, Germany) and serum neutralization assay as the gold standard. Moreover, a new commercial immunoassay, the PETIA test Nab (SGM, Rome, Italy), was used for neutralization evaluation. Statistical analysis was performed with R software, version 3.6.0. Results: Anti-SARS-CoV-2 IgG titers decayed during the first ninety days after the vaccine second dose. The following booster dose significantly (p < 0.001) increased IgG levels. A correlation between IgG expression and neutralizing activity modulation was found with a significant increase after the second and the third booster dose (p < 0.05. Compared to the Beta variant of the virus, the Omicron VOC was associated with a significantly larger quantity of IgG antibodies needed to achieve the same degree of neutralization. The best Nab test cutoff for high neutralization titer (≥1:80) was set for both Beta and Omicron variants. Conclusion: This study correlates vaccine-induced IgG expression and neutralizing activity using a new PETIA assay, suggesting its usefulness for SARS-CoV2 infection management.

Neutralizing Antibodies against SARS-CoV-2 Beta and Omicron Variants Inhibition Comparison after BNT162b2 mRNA Booster Doses with a New PETIA sVNT Assay.

BACKGROUND: Monitoring antibody response following SARS-CoV-2 vaccination is strategic, and neutralizing antibodies represent the gold standard. The neutralizing response to Beta and Omicron VOCs was evaluated versus the gold standard by a new commercial automated assay. METHODS: Serum samples from 100 healthcare workers from the Fondazione Policlinico Universitario Campus Biomedico and the Pescara Hospital were collected. IgG levels were determined by chemiluminescent immunoassay (Abbott Laboratories, Wiesbaden, Germany) and serum neutralization assay as the gold standard. Moreover, a new commercial immunoassay, the PETIA test Nab (SGM, Rome, Italy), was used for neutralization evaluation. Statistical analysis was performed with R software, version 3.6.0. RESULTS: Anti-SARS-CoV-2 IgG titers decayed during the first ninety days after the vaccine second dose. The following booster dose significantly (p < 0.001) increased IgG levels. A correlation between IgG expression and neutralizing activity modulation was found with a significant increase after the second and the third booster dose (p < 0.05. Compared to the Beta variant of the virus, the Omicron VOC was associated with a significantly larger quantity of IgG antibodies needed to achieve the same degree of neutralization. The best Nab test cutoff for high neutralization titer (≥1:80) was set for both Beta and Omicron variants. CONCLUSION: This study correlates vaccine-induced IgG expression and neutralizing activity using a new PETIA assay, suggesting its usefulness for SARS-CoV2 infection management.

Neutralization of SARS-CoV-2 Variants by Serum from BNT162b2 Vaccine Recipients.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has evolved rapidly, leading to viral lineages characterized by multiple mutations in the spike protein, which could potentially confer to the virus the ability to avoid the vaccine-induced immune response, making the vaccines less effective or ineffective. Here, we initially evaluated the neutralization capabilities in vitro by serum neutralization (SN) of six serum samples collected from recipients of the BNT162b2 vaccine against 11 SARS-CoV-2 isolates belonging to the major SARS-CoV-2 lineages that had been circulating in Italy. Then, we considered 30 additional serum samples by SN assay against the dominant B.1.617.2 (Delta) variant. A B.1 lineage isolate was used as a reference. In the first analysis, significant differences when compared with the reference strain (p > 0.05) were not evidenced; instead, when the panel of 30 sera was tested against the B.1.617.2 (Delta) variant, a significant (p = 0.0015) 2.38-fold reduction in neutralizing titres compared with the reference after the first vaccine dose was demonstrated. After the second vaccine dose, the reduction was not significant (p = 0.1835). This study highlights that the BNT162b2 vaccine stimulates a humoral response able to neutralize all tested SARS-CoV-2 variants, thus suggesting a prominent role in mitigating the impact of the SARS-CoV-2 pandemic in real-world conditions. Long-term follow-up is currently ongoing.

SARS-CoV-2 RNA Persistence in Naso-Pharyngeal Swabs.

Since February 2020, Italy has been seriously affected by the SARS-CoV-2 pandemic. To support the National Health Care system, naso-pharyngeal/oropharyngeal swabs collected from suspected cases of Teramo province, Abruzzo region, are tested at Istituto Zooprofilattico Sperimentale dell'Abruzzo e del Molise G. Caporale, for the presence of SARS-CoV-2 RNA. Out of 12,446 tested individuals, 605 returned positive results at least once, with prevalence significantly higher in men. A reduction of the level of viral RNA in the first swab per each positive patient collected over time was also observed. Moreover, 81 patients had at least one positive sample and two final negative tests: positivity in swabs lasted from 14 to 63 days, with a median value of 30 days. This shows the potential for the virus to coexist with patients for a long time, although we highlighted intermittent positivity in several cases. The evolution of the SARS-CoV-2 epidemiological situation and knowledge on viral shedding should be closely monitored, to interpret the findings correctly and adjust accordingly the surveillance activities.

Epidemiological Significance of SARS-CoV-2 RNA Dynamic in Naso-Pharyngeal Swabs.

From 16 March to 15 December 2020, 132,357 naso-pharyngeal/oropharyngeal swabs were collected in the province of Teramo, Abruzzo Region, Italy, and tested for the presence of SARS-CoV-2 genomic RNA by a commercially available molecular assay. A total of 12,880 swabs resulted positive. For 8212 positive patients (4.150 women and 4.062 men) the median age was statistically different between women (median: 49.55 ± 23.9 of SD) and men (median: 48.35 ± 23.5 of SD) while no differences were found in the comparison between the cycle threshold for the N protein-encoding gene (CT N) median values and gender. Differences were observed in the CT N gene median values of swabs collected from March to September as well as in the pairwise comparison between September and October and between November and December. The CT N gene median values observed in specific periods characterizing the SARS-CoV-2 epidemic in 2020 were also compared with the incidence of COVID-19 cases; a strong inverse correlation was highlighted (Pearson correlation coefficient = -0.978). Our findings confirm the usefulness of the CT N values as an indirect detection parameter to monitor viral loads in the population.

Novel SARS-CoV-2 Variants in Italy: The Role of Veterinary Public Health Institutes.

Novel SARS-CoV-2 variants with potential impacts on diagnostics, antivirals, and vaccines are spreading in Italy. In this editorial, we highlight the role that veterinary public health institutes may have in this global crisis, as their expertise in genomic/antigenic surveillance and animal studies are crucial to tackle SARS-CoV-2 pandemic.

Infection sustained by lineage B.1.1.7 of SARS-CoV-2 is characterised by longer persistence and higher viral RNA loads in nasopharyngeal swabs.

Following the announcement on December 2020 about the emergence of a new variant (VOC 202012/ 01, B.1.1.7 lineage) in the United Kingdom, a targeted surveillance was put in place in the Abruzzo region (Italy), which allowed detection of 313 persons affected by lineage B.1.1.7, up to the 20th of February 2021. We investigated the results of RT-PCR on nasopharyngeal swabs tested from December 2020 to February 2021 to verify any difference on the viral load and persistence between people infected by lineage B.1.1.7 and others. Statistically significant lower values of CT associated with the detection of the N protein encoding gene (CT N) were observed in persons with lineage B.1.1.7 infection (median CT N = 15.8)in comparison to those infected by other lineages (median CT N = 16.9). A significantly longer duration of the persistence of SARS-CoV-2 RNA in nasopharyngeal swabs was observed in persons with lineage B.1.1.7 infection (16 days) in comparison to those infected by other lineages (14 days).

A COVID-19 Hotspot Area: Activities and Epidemiological Findings.

By late March 2020, Villa Caldari, a small village of the municipality of Ortona (Abruzzo region), was registering an incidence rate of COVID-19 cases ten times greater than the overall municipality and was declared a hotspot area. Twenty-two days later, epidemiological investigation and sampling were performed, to evaluate SARS-CoV-2 circulation and the presence of SARS-CoV-2 antibodies. Overall, 681 nasopharyngeal swabs and 667 blood samples were collected. Only one resident of the village resulted in being positive for RNA viral shedding, while 73 were positive for SARS-CoV-2 antibodies. The overall seroprevalence was 10.9%. The difference between the seroprevalence of infection in asymptomatic and symptomatic individuals was significant (χ2 = 14.50 p-value = 0.0001). Amongst the residents positive for antibodies, fatigue and/or muscle pain, fever and anosmia were the most experienced symptoms, whose most frequent onset was observed during the first two weeks of March. Familial and habit-related clusters were highlighted. Nevertheless, the investigations showed a low SARS-CoV-2 circulation in the village at the time of the sampling, demonstrating virus transmission could be limited when strict emergency measures are followed. Given the favorable results, the emergency measures were then lifted.

SARS-CoV-2 replicates in respiratory ex vivo organ cultures of domestic ruminant species.

There is strong evidence that severe acute respiratory syndrome 2 virus (SARS-CoV-2), the causative agent of the coronavirus disease 2019 (COVID-19) pandemic, originated from an animal reservoir. However, the exact mechanisms of emergence, the host species involved, and the risk to domestic and agricultural animals are largely unknown. Some domestic animal species, including cats, ferrets, and minks, have been demonstrated to be susceptible to SARS-CoV-2 infection, while others, such as pigs and chickens, are not. Importantly, the susceptibility of ruminants to SARS-CoV-2 is unknown, even though they often live in close proximity to humans. We investigated the replication and tissue tropism of two different SARS-CoV-2 isolates in the respiratory tract of three farm animal species - cattle, sheep, and pigs - using respiratory ex vivo organ cultures (EVOCs). We demonstrate that the respiratory tissues of cattle and sheep, but not of pigs, sustain viral replication in vitro of both isolates and that SARS-CoV-2 is associated to ACE2-expressing cells of the respiratory tract of both ruminant species. Intriguingly, a SARS-CoV-2 isolate containing an amino acid substitution at site 614 of the spike protein (mutation D614G) replicated at higher magnitude in ex vivo tissues of both ruminant species, supporting previous results obtained using human cells. These results suggest that additional in vivo experiments involving several ruminant species are warranted to determine their potential role in the epidemiology of this virus.

A COVID-19 Hotspot Area: Activities and Epidemiological Findings

By late March 2020, Villa Caldari, a small village of the municipality of Ortona (Abruzzo region), was registering an incidence rate of COVID-19 cases ten times greater than the overall municipality and was declared a hotspot area. Twenty-two days later, epidemiological investigation and sampling were performed, to evaluate SARS-CoV-2 circulation and the presence of SARS-CoV-2 antibodies. Overall, 681 nasopharyngeal swabs and 667 blood samples were collected. Only one resident of the village resulted in being positive for RNA viral shedding, while 73 were positive for SARS-CoV-2 antibodies. The overall seroprevalence was 10.9%. The difference between the seroprevalence of infection in asymptomatic and symptomatic individuals was significant (χ2 = 14.50 p-value = 0.0001). Amongst the residents positive for antibodies, fatigue and/or muscle pain, fever and anosmia were the most experienced symptoms, whose most frequent onset was observed during the first two weeks of March. Familial and habit-related clusters were highlighted. Nevertheless, the investigations showed a low SARS-CoV-2 circulation in the village at the time of the sampling, demonstrating virus transmission could be limited when strict emergency measures are followed. Given the favorable results, the emergency measures were then lifted.

A "One-Health" approach for diagnosis and molecular characterization of SARS-CoV-2 in Italy.

The current pandemic is caused by a novel coronavirus (CoV) called SARS-CoV-2 (species Severe acute respiratory syndrome-related coronavirus, subgenus Sarbecovirus, genus Betacoronavirus, family Coronaviridae). In Italy, up to the 2nd of April 2020, overall 139,422 confirmed cases and 17,669 deaths have been notified, while 26,491 people have recovered. Besides the overloading of hospitals, another issue to face was the capacity to perform thousands of tests per day. In this perspective, to support the National Health Care System and to minimize the impact of this rapidly spreading virus, the Italian Ministry of Health involved the Istituti Zooprofilattici Sperimentali (IZSs), Veterinary Public Health Institutes, in the diagnosis of SARS-CoV-2 by testing human samples. The Istituto Zooprofilattico Sperimentale dell'Abruzzo e del Molise is currently testing more than 600 samples per day and performing whole genome sequencing from positive samples. Sequence analysis of these samples suggested that different viral variants may be circulating in Italy, and so in Abruzzo region. CoVs, and related diseases, are well known to veterinarians since decades. The experience that veterinarians operating within the Public Health system gained in the control and characterization of previous health issues of livestock and poultry including avian flu, bluetongue, foot and mouth disease, responsible for huge economic losses, is certainly of great help to minimize the impact of this global crisis.