Serological testing reveals the hidden COVID-19 burden among healthcare workers experiencing a SARS-CoV-2 nosocomial outbreak.

We describe the results of testing healthcare workers from a tertiary care hospital in Japan, which had experienced a COVID-19 outbreak during the first peak of the pandemic, for SARS-CoV-2 specific antibody seroconversion. Using two chemiluminescent immunoassays and a confirmatory surrogate virus neutralization test, serological testing unveiled that a surprising 42.2% (27/64) of overlooked COVID-19 diagnoses had occurred when case detection had relied solely on SARS-CoV-2 nucleic acid amplification testing. This undetected portion of the COVID-19 iceberg beneath the surface may potentially have led to silent transmissions and triggered the spread. A questionnaire-based risk assessment was further indicative of exposures to specific aerosol-generating procedures, i.e. non-invasive ventilation, having had conveyed the highest transmission risks and served as the origin of outbreak. Our observations are supportive of a multi-tiered testing approach, including the use of serological diagnostics, in order to accomplish exhaustive case detection along the whole COVID-19 spectrum.

Objective epizootic situation on cattle leukemia in Dagestan

Based on reliable factual material, the article presents an analysis of the objective epizootic situation on cattle leukemia that developed in 2020 in Dagestan Republic as a whole and in individual rural areas, urban districts and areas of distant pasture farming. The number of unfavorable points, cases of illness and the coefficient of focality are indicated. The level of infection of animals with BLV for the analyzed period was 1,35 %, the incidence rate - 17,43 %. At the same time, the value of the focal incidence indicator was 3,45. BLV infection is registered in most municipal districts and has an uneven distribution. According to epizootological data for the period under review, 11 administrative districts, the cities Dagestanskie Ogni, Derbent, Kizlyar and Bakres, Derbent, Ulankholsk zones of distant-pasture animal husbandry, are safe in terms of BLV infection. In 30 municipalities, the number of animals infected with the leukemia virus ranged from 0,10 to 3,00 %, in other 10 districts - from 3,25 to 7,51 %. At the beginning of 2020, there were 111 leukemia-unfavorable points in the region. During the year, 24 new foci were identified, 19 foci of infection were healed and by the end of the year 116 remained, including 38 - in the public sector, 78 - in the individual. The values on morbidity and ill-being are of an increasing nature. This situation is explained by the introduction of large-scale anti-leukemic measures of serological diagnostics, which made it possible to concentrate the implementation of health-improving measures in farms with the most widespread infection. In this situation, one of the ways to solve the problem is full coverage of diagnostic studies of animals, isolated rearing of young animals, exclusion of joint keeping, grazing and calving of healthy and infected animals, increasing the frequency of serological testing of seronegative animals until two consecutive negative results are obtained.

Systems serology detects functionally distinct coronavirus antibody features in children and elderly

The hallmarks of COVID-19 are higher pathogenicity and mortality in the elderly compared to children. Examining baseline SARS-CoV-2 cross-reactive immunological responses, induced by circulating human coronaviruses (hCoVs), is needed to understand such divergent clinical outcomes. Here we show analysis of coronavirus antibody responses of pre-pandemic healthy children (n = 89), adults (n = 98), elderly (n = 57), and COVID-19 patients (n = 50) by systems serology. Moderate levels of cross-reactive, but non-neutralizing, SARS-CoV-2 antibodies are detected in pre-pandemic healthy individuals. SARS-CoV-2 antigen-specific Fcγ receptor binding accurately distinguishes COVID-19 patients from healthy individuals, suggesting that SARS-CoV-2 infection induces qualitative changes to antibody Fc, enhancing Fcγ receptor engagement. Higher cross-reactive SARS-CoV-2 IgA and IgG are observed in healthy elderly, while healthy children display elevated SARS-CoV-2 IgM, suggesting that children have fewer hCoV exposures, resulting in less-experienced but more polyreactive humoral immunity. Age-dependent analysis of COVID-19 patients, confirms elevated class-switched antibodies in elderly, while children have stronger Fc responses which we demonstrate are functionally different. These insights will inform COVID-19 vaccination strategies, improved serological diagnostics and therapeutics.

First Evidence of Ehrlichia minasensis Infection in Horses from Brazil

The genus Ehrlichia includes tick-borne bacterial pathogens affecting humans, domestic and wild mammals. Ehrlichia minasensis has been identified in different animal species and geographical locations, suggesting that this is a widely distributed and generalist Ehrlichia. In the present study, we evaluated Ehrlichial infection in 148 Equidae presented to the Medical Clinic Department of a Veterinary Hospital from a midwestern region of Brazil. Blood samples and ticks collected from the animals were tested by Polymerase Chain Reaction (PCR) for the presence of Ehrlichia spp. A multigenic approach including Anaplasmataceae-specific (i.e., 16S rRNA, groEL, gltA) and Ehrlichia-specific (i.e., dsb and trp36) genes was used for accurate bacteria identification. Sera samples were also collected and evaluated for the detection of anti-Ehrlichia antibodies by indirect fluorescent antibody test (IFA). Possible associations between molecular and serological diagnostics and clinical and hematological manifestations were tested using chi-squared or Fisher’s exact tests. Sequence analysis of the dsb fragment revealed that three horses (2.03%) were exposed to E. minasensis. Sixty-one (41.2%) Equidae (58 equines and three mules), were seropositive for Ehrlichia spp., with antibody titers ranging between 40 and 2560. Seropositivity to ehrlichial antigens was statistically associated with tick infestation, rural origin, hypoalbuminemia and hyperproteinemia (p ≤ 0.05). The present study reports the first evidence of natural infection by E. minasensis in horses from Brazil.

Serological Diagnostics Of Atrophy Of The Gastric Mucosa

The association between HP infection and CAH development suggested by the Correa cascade is supported by several cohort studies. To diagnose and determine the severity of atrophy at the Center, patients underwent a serological analysis by ELISA method in the clinical and biochemical laboratory of the State Institution "RSNPMCT and MR". Analyzes were taken in 69 (58.5%) patients, including 35 (50.7%) with CAH, 34 (49.3%) with CNG. Analysis of the non-invasive indicator of the level of pepsinogens in patients with CAH determined that severe atrophy was detected in 20%, moderate atrophy - 42.9%, weak atrophy - 34.3%, and no atrophy was found in 2.9% of patients. The pepsinogen parameters in patients with CNG were also as follows: PG I was within 95.4 ± 7.2 μg / L, PG II 14.5 ± 1.3 and PGI / PGII 6.6 ± 0.2. Pepsinogen values were more pronounced in patients with CAH: with a pronounced degree of atrophy, PGI was 8.7 ± 0.1 μg / l and PGI / PGII was 1.1 ± 0.1; with moderate atrophy 16.6 ± 0.9 μg / l and 1.6 ± 0.1; with mild atrophy 27.2 ± 1.5 and 2.3 ± 0.2, respectively.

Study of the activity and specifity of microseries of Yersinia Enterocolitica sera O3, O5, O6.30, O8, O9 in vitro SAT

Yersiniosis is ubiquitous and causes great damage and death. The genus Yersinia has 11 species, 3 of which are characteristic of human disease: Yersinia pestis, Yersinia enterocolitica and Yersinia pseudotuberculosis. Based on serotyping, this pathogen is classified into more than 57 serogroups. Pathogenic serotypes are O: 3 (biogroup 4), O: 5.27 (biogroup 2 and 3), O: 8 (biogroup 1B) and O: 9 (biogroup 2). The most common serogroups isolated from humans in European countries are O: 3 and then O: 9. In the United States, for example, the more common serogroup O: 8. In general, 11 serovars are most associated with human infections (O: 3, O: 4, O: 5.27, O: 8, O: 9, O: 13, O: 18, O: 20, O: 21). The publication presents data on the activity of two microseries of Yersinia Enterocolitica serovariants O3, O5, O6.30, O8, O9 by serological predictions in the samples of RA. In the process of obtaining and increasing the activity of two microseries of microseries, the methodology of obtaining components in a set, designated for serological diagnostics of tincture in the RA, was developed. The effectiveness of experimental samples of Yersinia sera was studied. Serum Yersinia enterocolitica O3 of microseries 1, O8 of microseries 2 and O9 of both microseries were determined to be highly specific. Yersinia sera of serovar O6.30 microseries 1, as well as O3, O5, O9 of both microseries are active in a titer of not less than 1: 400. It is established that according to this method of obtaining RA components allows to detect specific antibodies in the sera of animals in the titer of 1: 400–1: 800. Due to the antigenic affinity of strains of serotypes O5 and O6.30, there is a need to pay more attention to the selection of Yersinia, in particular to work with antigenically homogeneous clones, which will avoid nonspecific intraspecific cross-reactions, respectively, increase the specificity of sera.

Structure-based design of prefusion-stabilized SARS-CoV-2 spikes

The coronavirus disease 2019 (COVID-19) pandemic has led to accelerated efforts to develop therapeutics and vaccines. A key target of these efforts is the spike (S) protein, which is metastable and difficult to produce recombinantly. We characterized 100 structure-guided spike designs and identified 26 individual substitutions that increased protein yields and stability. Testing combinations of beneficial substitutions resulted in the identification of HexaPro, a variant with six beneficial proline substitutions exhibiting higher expression than its parental construct (by a factor of 10) as well as the ability to withstand heat stress, storage at room temperature, and three freeze-thaw cycles. A cryo–electron microscopy structure of HexaPro at a resolution of 3.2 angstroms confirmed that it retains the prefusion spike conformation. High-yield production of a stabilized prefusion spike protein will accelerate the development of vaccines and serological diagnostics for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2).

Stabilizing the Closed SARS-CoV-2 Spike Trimer

The trimeric spike (S) protein of SARS-CoV-2 is the primary focus of most vaccine design and development efforts. Due to intrinsic instability typical of class I fusion proteins, S tends to prematurely refold to the post-fusion conformation, compromising immunogenic properties and prefusion trimer yields. To support ongoing vaccine development efforts, we report the structure-based design of soluble S trimers, with increased yields and stabilities, based on introduction of single point mutations and disulfide-bridges. We identify two regions in the S-protein critical for the protein’s stability: the heptad repeat region 1 of the S2 subunit and subunit domain 1 at the interface with S2. We combined a minimal selection of mostly interprotomeric mutations to create a stable S-closed variant with a 6.4-fold higher expression than the parental construct while no longer containing a heterologous trimerization domain. The cryo-EM structure reveals a correctly folded, predominantly closed pre-fusion conformation. Highly stable and well producing S protein and the increased understanding of S protein structure will support vaccine development and serological diagnostics.

Structure-based Design of Prefusion-stabilized SARS-CoV-2 Spikes

ABSTRACTThe COVID-19 pandemic caused by the novel coronavirus SARS-CoV-2 has led to accelerated efforts to develop therapeutics, diagnostics, and vaccines to mitigate this public health emergency. A key target of these efforts is the spike (S) protein, a large trimeric class I fusion protein that is metastable and difficult to produce recombinantly in large quantities. Here, we designed and expressed over 100 structure-guided spike variants based upon a previously determined cryo-EM structure of the prefusion SARS-CoV-2 spike. Biochemical, biophysical and structural characterization of these variants identified numerous individual substitutions that increased protein yields and stability. The best variant, HexaPro, has six beneficial proline substitutions leading to ∼10-fold higher expression than its parental construct and is able to withstand heat stress, storage at room temperature, and multiple freeze-thaws. A 3.2 Å-resolution cryo-EM structure of HexaPro confirmed that it retains the prefusion spike conformation. High-yield production of a stabilized prefusion spike protein will accelerate the development of vaccines and serological diagnostics for SARS-CoV-2.

CASES OF ASCARIASIS IN THE KRASNODAR REGION IN THE PERIOD FROM 2015 TO 2019

To identify the epidemiological situation in the Krasnodar region, the following activities were carried out: a) analysis of statistical data kindly provided to us by the Office of the Federal state statistics Service for the Krasnodar region in the last five years (2015–2019); b) analysis of the results of coprological studies of faecal smears; as well as the results of blood serum tests for serological diagnostics of children and adults, conducted in the laboratories of the GBUZ regional hospital No. 2 and KBUZ Research Institute – Regional clinical hospital No. 1, as well as in the laboratory of the emergency care hospital of Krasnodar city. Based on our research, we identified two categories of the population, of different age groups, most susceptible to the disease ascariasis. The first group is children 7–16 years old, students of Junior and senior classes of city schools and the second group is adults – from 18 years and older, students of colleges, technical schools, and working people. In the first group, the peak of infestation of children with ascariasis occurred in 2018, it reached 23.1% per 100,000 people surveyed. In 2019, it was lower than 22.8%, but still quite large. In the second group, the rise of the disease occurred in 2016, it amounted to 17.5%, in subsequent years, the decline of ascariasis is noticeable.