Six splice site variations, three of them novel, in the ABO gene occurring in nine individuals with ABO subtypes

Background Nucleotide mutations in the ABO gene may reduce the activity of glycosyltransferase, resulting in lower levels of A or B antigen expression in red blood cells. Six known splice sites have been identified according to the database of red cell immunogenetics and the blood group terminology of the International Society of Blood Transfusion. Here, we describe six distinct splice site variants in individuals with ABO subtypes. Methods The ABO phenotype was examined using a conventional serological method. A polymerase chain reaction sequence-based typing method was used to examine the whole coding sequence of the ABO gene. The ABO gene haplotypes were studied using allele-specific primer amplification or cloning technology. In silico analytic tools were used to assess the functional effect of splice site variations. Results Six distinct variants in the ABO gene splice sites were identified in nine individuals with ABO subtypes, including c.28 + 1_2delGT, c.28 + 5G > A, c.28 + 5G > C, c.155 + 5G > A, c.204-1G > A and c.374 + 5G > A. c.28 + 1_2delGT was detected in an Aw individual, while c.28 + 5G > A, c.28 + 5G > C, and c.204-1G > A were detected in Bel individuals. c.155 + 5G > A was detected in one B3 and two AB3 individuals, whereas c.374 + 5G > A was identified in two Ael individuals. Three novel splice site variants (c.28 + 1_2delGT, c.28 + 5G > A and c.28 + 5G > C) in the ABO gene were discovered, all of which resulted in low antigen expression. In silico analysis revealed that all variants had the potential to alter splice transcripts. Conclusions Three novel splice site variations in the ABO gene were identified in Chinese individuals, resulting in decreased A or B antigen expression and the formation of ABO subtypes.

Both the serum AFP test and AFP/GPC3/SALL4 immunohistochemistry are beneficial for predicting the prognosis of gastric adenocarcinoma

Background Both gastric adenocarcinoma with primitive enterocyte phenotype (GAPEP) (including hepatoid adenocarcinoma) and alpha-fetoprotein (AFP)-producing gastric adenocarcinoma have poor prognoses. However, the value of the serum AFP test and AFP/glypican-3 (GPC3)/spalt-like transcription factor 4 (SALL4) immunohistochemistry is still not clear, and these two methods have not yet been thoroughly compared. Methods We collected 421 consecutive non-neoadjuvant surgically or endoscopically resected gastric adenocarcinoma patients with serum AFP results before surgery (group A). We divided these cases into serum AFP-high (sAFP-H) and serum AFP-normal (sAFP-N) by serum AFP levels, and into GAPEP (expressing AFP, GPC3, or SALL4) and non-GAPEP (nGAPEP) by AFP/GPC3/SALL4 immunohistochemistry results. We also collected 12 non-resected gastric adenocarcinoma patients with serum AFP ≥ 7 ng/mL before treatment (group B). We analyzed these patients’ clinicopathological characteristics and prognoses. Results Seventeen (4.04%) patients in group A were sAFP-H. These patients were younger and mainly had tubular adenocarcinoma with later pT (P = 0.014) and pN (P = 0.047) categories and more lymphovascular invasion (P < 0.001), perineural spread (P = 0.008), and metastases or recurrence (P < 0.001). For immunohistochemistry, 34 (8.08%) cases were GAPEP, and GAPEP cases also had later pT categories than nGAPEP cases (P = 0.001). Most group B patients with elevated serum AFP (especially > 1000 ng/mL) had simultaneous metastases, mainly liver metastases. Both the serological method and immunohistochemical method were useful for predicting prognosis (AUC sAFP = 0.625, AUC A/G/S-IHC = 0.723, z statistic = 1.726, P = 0.084). The serum AFP level (especially > 1000 ng/mL) is more specific (100%), and immunohistochemistry is more sensitive (50%). Conclusion Both the serum AFP level and immunohistochemical expression of AFP/GPC3/SALL4 can be used to indicate a poor prognosis for gastric adenocarcinoma.

Combined use of two rapid tests for the conclusive diagnosis of Chagas disease: a systematic scoping review

ObjectiveThe goal of this systematic scoping review is to collect and summarise scientific evidence regarding the validity of two simultaneous immunochromatographic tests for the conclusive diagnosis of Chagas disease. The research was informed by the following review questions: Will the use of two rapid tests be a valid method for the definitive diagnosis of Chagas disease when compared with conventional serological tests? In what type of population has the operation of two rapid tests been tried for the diagnosis of Chagas disease? What are the biomedical and public health advantages of the diagnostic method resulting from the combination of two rapid tests over the conventional serological method? Will it be a cost–benefit strategy for the diagnosis of Chagas with respect to conventional serological tests?DesignSystematic scoping review.SettingA search of the published and unpublished literature in five databases was carried out, in order to identify, screen and select the studies included in this review.Results468 studies were identified, of which 46 were screened with a full-text reading, and finally, three articles were included in the review. All studies were in endemic countries with adult and paediatric populations (n=1133) and, together, they evaluated four different rapid tests. The rapid tests showed good sensitivity (97.4%–100%) and specificity (96.1%–100%) for the diagnosis of Chagas when used in combination and compared with the reference tests.ConclusionsThe simultaneous use of at least two immunochromatographic rapid tests is a valid option for the definitive diagnosis of chronic Chagas in endemic rural areas, as long as there are studies that previously evaluate their performance on the areas of implementation. Therefore, this could be an alternative to the current diagnostic standard. However, additional studies are still needed in more countries in order to provide further evidence and to investigate the cost–benefit.

An Enzyme-Linked Immunosorbent Assay for the Detection of Antibodies to Linguatula serrata in Experimentally Infected Dogs

Background: Linguatula serrata is a causative agent of visceral and nasopharyngeal linguatulosis in humans and animals. The aim of the present study was to investigate the immune response of dogs experimentally infected by L. serrata with ELISA. Methods: Five puppies were infected by inserting the L. serrata nymphs in their nasal cavities (infected group) in the Department of Parasitology of Shahid Chamran University of Ahvaz, during 2018-2019. Three animals were kept as the non-infected control group. Blood samples were collected from the animals for seven months at approximately monthly intervals for serum preparation. Nasal samples were taken weekly from the fourth month. ELISA was designed and performed on 64 sera (24 negatives, and 40 positives) using somatic (S), and excretory-secretory (ES) antigens. Results: Overall, 100% of the animals were infected with the parasite. Based on the results of ELISA, the ES antigen (sensitivity 95% and specificity 92%) was more preferred than the S antigen (sensitivity 95% and specificity 85%). Female parasites had significant effects on the immune response. There was a significant correlation between the clinical symptoms and the presence of female parasites (P<0.05). Conclusion: The results showed a practical method for dogs' experimental infection. ELISA method is suitable for the detection of infection at different stages of development, especially before the maturation stage of the parasite. In this regard, the ES antigen of the parasite was more immunogenic. Therefore, ELISA can be used as a serological method in the early detection and epidemiological studies of infection with L. serrata in dogs.

Helicobacter pylori genotypes in patients with stable angina combined with chronic gastritis

Research objective. То study the genotypes of Helicobacter pylori and their antibiotic sensitivity in patients with stable angina in combination with chronic gastritis. Material and methods. 46 patients with stable angina with a combination of chronic H. pylori-associated gastritis were included in the open prospective clinical study. To diagnose H. pylori, serological method of detection of antibodies in blood serum was used, PCR - diagnosis of H. pylori genes. Antibiotic sensitivity of H. pylori strains was studied by serial dilution method. Results. In the patients (n = 46) stable angina in combination with chronic gastritis in the endoscopic study by the EGDS method, gastrobioptates were obtained and further investigated. Chronic neatrophic gastritis was diagnosed in 54.3% of patients, atrophic gastritis - in 45.7%. It was found that the genotypes of H. pylori VacA had 8.7% of patients, CagA - 34.7%, HopQ - 13.1%, Oip - 30.4% of patients. Only 13.1% of patients had non-toxic genotypes. The absence of antibiotic resistance of the first line of erication therapy - clarithromycin and amoxicillin - was revealed. 45.7% of patients showed resistance, 39.1% showed weak sensitivity of H. pylori isolates to metronidazole. Conclusions. In patients with stable angina with a combination of chronic gastritis, H. pylori strains with toxigenic genotypes: CagA, Oip, Vac A, HopQ predominate. Resistance of H. pylori isolates to metronidazole was determined in 45.7% of patients.

Serological and molecular survey of Trypanosoma evansi in Camels (Camelus dromedarius) in Maiduguri, North-east, Nigeria

To date, camels still remain an important work animal as well as source of protein to humans in the Sudan and Sahel regions of Nigeria. Therefore, a cross-sectional study was conducted on 150 camels slaughtered in Maiduguri central abattoir to determine the prevalence of Trypanosoma evansi using Card Agglutination Test (CATT) and Polymerase Chain Reaction (PCR) techniques. Overall, 30 (20%) of the camels tested were seropositive while PCR targeting the 227 base pair of the Variable Surface Glycoprotein (VSG) gene of T. evansi detected the DNA of the parasite in 9 out of the 30seropositive camels. Higher infection was found among adult compared to the young camels using the two diagnostic techniques; 24.1% vs 19.0% and 10.3% vs 4.6%, for CATT and PCR techniques, respectively. However, the differences being not statistically significant (P > 0.05) for the two methods of diagnosis. Furthermore, significantly (P < 0.05)higher prevalence of infection was recorded among male compared to female camels using the serological method of diagnosis, while (P > 0.05) using the molecular method; 27.5% vs 13.6% for CATT and 10.1% vs 2.5% for PCR. Camels with PCV =24 %( mean: 19.8923 ± 4.0931) recorded significantly (P < 0.05) higher prevalence of 23.1% than those with PCV = 25% (mean 31.7294 ± 5.50584), where the prevalence was 17.6%.The results of this study showed that camel trypanosomosis is endemic in the study area. Furtherstudiesto elucidate the epidemiology and socioeconomic impact of this disease in the northeast region of Nigeria are desirable. Keywords:Serology, PCR, Dromedary camel, T.evansi, Maiduguri

A Novel Molecular Method for Simultaneous Identification of Vibrio parahaemolyticus 57 K-Serogroups Using Probe Melting Curve Analysis

The serotyping of Vibrio parahaemolyticus, which is crucial to the surveillance and detection of outbreaks of vibriosis infection, has been widely used in many countries. In this study, we developed a molecular assay, named multiplex ligation reaction based on probe melting curve analysis (MLMA), for simultaneous identification of V. parahaemolyticus 57 K-serogroups. Based on the previous genomes of 418 strains including 39 K-serogroups and the 18 K-serogroups sequences from public databases, we obtained 57 K-serogroups specific gene sequences for designing primers and probes. The developed MLMA assay for identifying the V. parahaemolyticus 57 K-serogroups showed high reproducibility, with the intra- and inter-assay standard deviations and coefficients of variation of no more than 1°C and 1%, respectively. The limit of detection for all gene targets ranged from 0.1 to 1.0 ng/µl. We validated the MLMA assay with a double-blind test identifying 595 V. parahaemolyticus isolates using conventional serotyping methods for comparison. The results showed the kappa value between the MLMA assay and the traditional serological method was 0.936 and that there was a 96.97% consistency rate with conventional serotyping methods for all detected isolates. Additionally, five rare K-serogroups were identified using the MLMA assay, as well as 18 strains that could not be identified using the traditional serotyping method. Thus, the MLMA assay provides a rapid, robust, and promising tool for the molecular serotyping of V. parahaemolyticus K-serogroups and has the potential application to the detection of outbreaks and surveillance of V. parahaemolyticus infection.

ŞEROLOGICAL EVIDENCE OF INFECTIOUS BURSAL DISEASE VIRUS INFECTION IN GREY BREASTED HELMET GUINEA FOWLS (Numida meleagris galeata Pallas) IN AN ARID ZONE OF NIGERIA

A survey of guineafowl sera from Maiduguri area of Borno State for antibody to infectious bursal disease (IBD) virus was carried out between the months of January to April, 1991. Agar gel diffusion precipitation test (AGDT) was the serological method employed for the investigation. Out of the total of 196 sera tested, only 16 were found to contain precipitating antibodies to IBD. This represents 9.1% infection rate.

HIV-1 Accessory Proteins: Which one is Potentially Effective in Diagnosis and Vaccine Development?

Background:: The combination antiretroviral therapy (cART) could increase the number of circulating naive CD4 T lymphocytes, but was not able to eradicate human immunodeficiency virus-1 (HIV-1) infection. Objective:: Thus, induction of strong immune responses is important for control of HIV-1 infection. Furthermore, a simple and perfect serological method is required to detect virus in untreated-, treated- and drug resistant- HIV-1 infected individuals. Methods:: This study was conducted to assess and compare immunogenic properties of Nef, Vif, Vpr and Vpu accessory proteins as an antigen candidate in mice and their diagnostic importance in human as a biomarker. Results:: Our data showed that in mice, all heterologous prime/ boost regimens were more potent than homologous prime/ boost regimens in eliciting Th1 response and Granzyme B secretion as CTL activity. Moreover, the Nef, Vpu and Vif proteins could significantly increase Th1 immune response. In contrast, the Vpr protein could considerably induce Th2 immune response. On the other hand, among four accessory proteins, HIV-1 Vpu could significantly detect treated group from untreated group as a possible biomarker in human. Conclusion:: Generally, among accessory proteins, Nef, Vpu and Vif antigens were potentially more suitable vaccine antigen candidates than Vpr antigen. Human antibodies against all these proteins were higher in HIV-1 different groups than healthy group. Among them, Vpu was known as a potent antigen in diagnosis of treated from untreated individuals. The potency of accessory proteins as an antigen candidate in an animal model and a human cohort study are underway.

The acute cholecystitis in COVID-19 patients: treatment in conditions of reprofiled hospital

Introduction. Routine surgical care has been suspended during the COVID-19 pandemic. For the treatment of patients with acute cholecystitis, conservative treatment, percutaneous drainage of the gallbladder or cholecystectomy is offered. Tactics of treatment of patients with acute cholecystitis against the background of COVID-19 have not been studied. It is important to study the data concerning the time of cholecystectomy in acute cholecystitis, comparing “early” and “delayed” cholecystectomy, which is performed after a period of conservative therapy. Aim. To present and evaluate the results of treatment of patients with acute cholecystitis against the background of COVID-19 in the conditions of a repurposed multi-specialty hospital on the basis of the O. M. Filatov Clinic Hospital No. 15 in Moscow. Material and methods. A retrospective comparative study with history control included 16 patients with acute cholecystitis against the background of COVID-19. Mechanical jaundice syndrome was diagnosed in 3 (18.75%) patients. The diagnosis of coronavirus infection using PCR was confirmed in 5 patients, serological method-in 2 patients, and in 9 patients the diagnosis was confirmed by X-ray or CT examination with negative/doubtful PCR test results. Results. Nine (56.25%) patients were operated 4 (25%) percutaneous interventions were performed, in 3 (18.75%) cases conservative therapy was performed. Most patients were operated on within the first day of admission. In the main group, a fatal outcome occurred in 1 (6.25%) case (death from a thromboembolic complication in a patient with mechanical jaundice syndrome). Discussion. The choice of «early» cholecystectomy for acute cholecystitis, provided the condition is stable and the initial changes in the lungs are appropriate in most COVID-19 patients. Conclusion. Performing cholecystectomy in patients with coronavirus infection in an infectious hospital did not lead to an increase in the duration of inpatient treatment and the prevalence of complications. The approaches mentiobed in the results section can be used as a safe method in the discussed category of patients.