PCR-positive Samples Research Articles

Comparison of L452R mutation variant diagnosis in SARS-COV-2 PCR positive samples with two different qPCR kits

Objectives: Quantitative reverse transcription‐polymerase chain reaction (qPCR) is used as the gold standard method to diagnose COVID-19 infection caused by SARS-CoV-2 which is the cause of the most important epidemic in world history. It was aimed to compare the results of two of the most commonly used commercial kits for the diagnosis of SARS-CoV-2 mutation in our laboratory during the pandemic. Methods: Our study included 5000 SARS-CoV-2 PCR positive nasopharyngeal swab samples (2500 L452R mutation positive samples, 2500 L452R mutation negative samples). PCR positivity and negativity of the L452R mutation of the positive SARS-CoV-2 positive samples were identified with the Diagnovital® (DIAGNO5plex NS SARS-CoV-2 Real Time PCR Kit [A1 Life Sciences Istanbul]) kit. The mentioned samples were also studied with a different commercial PCR kit, Bio-Speedy® (SARS-CoV-2 Emerging Plus Real Time PCR Kit [Bioeksen R&D Technologies Istanbul]). Results: A total of 5000 samples included in the study were concluded as SARS-CoV-2 positive with both tests. One hundred and fifty of 2500 samples that were found positive for SARS-CoV-2 but negative for L452R mutations with the Diagnovital® kit were found positive with the Bio-Speedy® kit for SARS-CoV-2. The compatability between the two kits was found to be high (Kappa= 0.940). The mean Ct values of the samples found positive with the Diagnovital® kit and Bio-Speedy® kit were 24.15 ± 6.75 and 20.72 ± 7.17, respectively and the difference was statistically significant. Conclusions: It was determined the two commercial kits included in the study were extremely compatible based on their analysis. Therefore both kits can be used safely for COVID-19 symptomatic patients.

Rapid point-of-care detection of SARS-CoV-2 infection in exhaled breath using ion mobility spectrometry: a pilot study

BackgroundAn effective testing strategy is essential for pandemic control of the novel Coronavirus disease 2019 (COVID-19) caused by infection with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Breath gas analysis can expand the available toolbox for diagnostic tests by using a rapid, cost-beneficial, high-throughput point-of-care test. We conducted a bi-center clinical pilot study in Germany to evaluate breath gas analysis using multi-capillary column ion mobility spectrometry (MCC-IMS) to detect SARS-CoV-2 infection.MethodsBetween September 23, 2020, and June 11, 2021, breath gas measurements were performed on 380 patients (SARS-CoV-2 real-time polymerase chain reaction (PCR) positive: 186; PCR negative: 194) presenting to the emergency department (ED) with respiratory symptoms.ResultsBreath gas analysis using MCC-IMS identified 110 peaks; 54 showed statistically significant differences in peak intensity between the SARS-CoV-2 PCR-negative and PCR-positive groups. A decision tree analysis classification resulted in a sensitivity of 83% and specificity of 86%, but limited robustness to dataset changes. Modest values for the sensitivity (74%) and specificity (52%) were obtained using linear discriminant analysis. A systematic search for peaks led to a sensitivity of 77% and specificity of 67%; however, validation by transferability to other data is questionable.ConclusionsDespite identifying several peaks by MCC-IMS with significant differences in peak intensity between PCR-negative and PCR-positive samples, finding a classification system that allows reliable differentiation between the two groups proved to be difficult. However, with some modifications to the setup, breath gas analysis using MCC-IMS may be a useful diagnostic toolbox for SARS-CoV-2 infection.Trial registration: This study was registered at ClinicalTrials.gov on September 21, 2020 (NCT04556318; Study-ID: HC-N-H-2004).

A seromolecular study to determine the prevalence of cytomegalovirus in pregnant women referred to health centers in the north of Iran.

Because of the controversial aspects of the CMV virus during pregnancy, it should be considered a serious health threat, especially in developing countries. The present seromolecular study aimed to determine cytomegalovirus prevalence in pregnant women referred to health centers in the north of Iran. One hundred and twenty-five pregnant women who were referred to health centers in Mazandaran province for regular health checks were randomly selected from Jan 2022 to Oct 2022. To detect the presence of the CMV genome and specific IgM and IgG antibodies against cytomegalovirus, the conventional PCR and ELISA tests were applied respectively. All 125 pregnant women that attended the study were from Mazandaran province with a mean age of 30 years ranging from 20 to 42 years. The result showed that 2 (1.6%), 92 (73.6%), and 2 (1.6%) of the cases were positive for IgM, IgG, and IgM/IgG, respectively. The PCR test results indicated that the CMV DNA was present in 10 (8%) pregnant women. Our study shows that all PCR-positive cases were negative for the IgM test. Of the 10 PCR-positive samples 3 were positive and 1 was suspicious for the IgG test. Our study revealed that there is an urgent need for vaccination or other strategies to prevent and treat congenital CMV infection. Reducing the burden of congenital CMV infection requires global awareness. Further studies are recommended to obtain accurate estimates of the risk of congenital CMV infection.

Molecular Prevalence Study of QX-like Infectious Bronchitis Virus in Japan from 2016 to 2023.

Infectious bronchitis is an acute and highly contagious disease in chickens caused by the infectious bronchitis virus (IBV), which has caused significant economic losses to the poultry industry worldwide. The antigenic variant, the QX-like genotype (GI-19 lineage), has been currently reported in epidemics in East Asia, Southeast Asia, the Middle East, Europe, and Africa. We first reported an epidemic of Japanese QX-like IBVs genetically related to Chinese and South Korean strains in the Kyushu region of Japan in 2020. However, because their nationwide prevalence was unknown, we performed a nationwide survey. The testing of 419 reverse transcription (RT)-PCR-positive samples (376 layers and 43 broilers) of IBV field strains between April 2016 and March 2022 detected two QX-like IBVs in 2019 and 2021 broiler samples from one region. A survey of fecal samples collected from 122-layer farms nationwide between November 2022 and January 2023 detected QX-like IBV genes from seven farms in various regions. Phylogenetic tree analysis on the basis of the S1 gene showed that all QX-like IBVs detected in Japan were genetically related to recent Chinese and South Korean strains. A new RT-PCR assay was developed to distinguish between QX-like IBV and other IBV variants prevalent in Japan, whose results were consistent with those of previously reported identification methods. These results suggest that QX-like IBV is rapidly spreading in Japan and that countermeasures are necessary.

Assessing the effectiveness of sample and RNA extract pool testing for the detection of SARS-CoV-2 RNA by real time RT-PCR

Introduction: Pooled testing is a cost-effective approach to increasing testing capacity during pandemics. This study analysed the effectiveness of pooling of samples and RNA extracts for the detection of SARS-CoV-2 by real time RT-PCR.Methods: Twenty SARS-CoV-2 positive samples with Ct value of 25–35 and 60 known negative samples based on initial PCR results were used for this study. The samples were used to prepare 2-, 4- and 8-fold pooled samples prior to extraction. The RNA extracts of a further 10 PCR positive samples were pooled to prepare 2-, 4- and 8-fold RNA extract pools. The Ct values of neat samples and pooled samples were compared using the paired t-test with a 95% confidence interval. The same was done for the neat RNA extracts and the extract pools.Results: The detection capacity was considerably lower when the pool size was increased from 2- to 8-fold in sample pooling, whereas pools of RNA extracts showed 100% detection from 2- to 8-fold dilution. The increase in Ct value of 2-, 4- and 8-fold sample dilutions were 1.69 ± 0.78, 3.84 ± 1.47 and 8.98 ± 2.25, respectively (P < 0.0001). However, there was a small rise in the Ct value of 2-, 4- and 8-fold extract pools (1.01 ± 0.38, 2.18 ± 0.82 and 3.05 ± 0.77, respectively).Conclusions: Large scale screening of asymptomatic individuals for SARS-CoV-2 can be maximised with optimal use of resources by 2- or 4-fold pooling of samples or 4- or 8-fold pooling of RNA extracts without significantly compromising the detection capacity.

Plasmodium falciparum with pfhrp2/3 Deletion Not Detected in a 2018-2021 Malaria Longitudinal Cohort Study in Kinshasa Province, Democratic Republic of the Congo.

Histidine-rich protein 2- (HRP2-) based rapid diagnostic tests (RDTs) are widely used to detect Plasmodium falciparum in sub-Saharan Africa. Reports of parasites with pfhrp2 and/or pfhrp3 (pfhrp2/3) gene deletions in Africa raise concerns about the long-term viability of HRP2-based RDTs. We evaluated changes in pfhrp2/3 deletion prevalence over time using a 2018-2021 longitudinal study of 1,635 enrolled individuals in Kinshasa Province, Democratic Republic of the Congo (DRC). Samples collected during biannual household visits with ≥ 100 parasites/µL by quantitative real-time polymerase chain reaction were genotyped using a multiplex real-time PCR assay. Among 2,726 P. falciparum PCR-positive samples collected from 993 participants during the study period, 1,267 (46.5%) were genotyped. No pfhrp2/3 deletions or mixed pfhrp2/3-intact and -deleted infections were identified in our study. Pfhrp2/3-deleted parasites were not detected in Kinshasa Province; ongoing use of HRP2-based RDTs is appropriate.

Molecular detection and pyrG sequence analysis of Avibacterium paragallinarum using clinical samples of infraorbital exudates from layer chickens with infectious coryza symptoms in Indonesia.

Infectious coryza (IC) or snot, is caused by Avibacterium paragallinarum and leads to upper respiratory disease in poultry. Various diagnostic methods are available, including isolation and identification through bacterial culture and biochemical tests. However, the isolation and subsequent identification of A. paragallinarum are challenging because the bacteria are fastidious and require specific growth factors. This study aimed to detect A. paragallinarum in clinical samples taken from the exudate of the infraorbital sinus of layer hens showing clinical signs of IC. Samples were collected from 10 layer hens with IC symptoms. Following DNA extraction, HPG-2 polymerase chain reaction (PCR) assays were performed. The PCR amplicons underwent electrophoresis to determine those of the correct target size (511 bp), and these were sequenced. The resultant sequences were analyzed using the National Center for Biotechnology Information (NCBI) basic local alignment search tool. MEGA X was used for bioinformatics analysis. The presence of A. paragallinarum was confirmed by HPG-2 PCR in 4/10 samples. Bioinformatics analysis showed that the amino acid sequence of the samples and the A. paragallinarum reference sequences in the NCBI database were grouped within the same cluster. Furthermore, the nucleotide sequences showed 98.64%-100% of similarity with the reference sequences. The phylogenetic reconstruction of partial pyrG sequences from 55 A. paragallinarum strains/isolates deposited in GenBank confirmed that the four HPG-2 PCR-positive samples fell within the A. paragallinarum cluster, separate from the Pasteurella multocida, Avibacterium spp., and Rodentibacter pneumotropicus clusters. Avibacterium paragallinarum infection was molecularly confirmed in 4/10 (40%) samples by HPG-2 PCR amplicon detection. Clustering of the pyrG partial gene sequences revealed that the positive samples fell within the A. paragallinarum cluster.

Molecular detection and phylogenetic analysis of Toxocara vitulorum in feces and milk samples from naturally infected water buffaloes

Toxocara vitulorum infects cattle and water buffalo, leading to mild to severe infection in calves and has wide geographic distributions, especially in tropical and subtropical countries. This work aimed to assess the prevalence, distributions, and phylogeny of T.vitulorum in water buffaloes in different parts of Sivas, one of the essential buffalo-breeding areas in Türkiye. T.vitulorum was found in 42 (8.23%) and 54 (10.58%) fecal (n:510) samples using microscopic and molecular techniques, respectively. T.vitulorum was higher in animals aged 0–6 months compared to other groups. Furthermore, when animals aged 0–6 months were grouped within themselves, the prevalence of T.vitulorum in 1–3 month-old-animals was higher than in both younger than one month and older than three months. T.vitulorum was detected in fecal samples obtained from animals older than six months. In colostrum/milk samples (n:100), T.vitulorum-larvae were found in 4% and 10% with microscopic and molecular techniques, respectively. The larvae were detected in colostrum/milk samples in the mother between the 2nd and 28th days postpartum-period. The ITS-1-gene of 11 PCR-positive samples was sequenced. The 98.99–100% nucleotide identity was determined between our T.vitulorum isolates and those present in GenBank.In conclusion, this is the first molecular survey and phylogenetic analysis of T.vitulorum in fecal and colostrum/milk samples from naturally infected water buffaloes. Data obtained in this study will help to understand the life cycle and epidemiology of the nematode. Data also revealed that veterinarians should consider older animals as well as young animals in their control program of nematode infections in farms.

Comparison of COVID-19 Rapid Antigen Test Results with PCR Reactive/ Non-Reactive Cases

Objective: To evaluate the analytical accuracy of the COVID-19 Ag Rapid Test Device by Abbott PanbioTM using RT-PCR as the reference assay and to detect any false-positive/negative reactions to assess the specificity of this analytical procedure.
 Study Design: Cross-Sectional analytical study.
 Place and Duration of Study: Department of Chemical Pathology, Chughtai Institute of Pathology, Lahore Pakistan, from Feb to May 2021.
 Methodology: This study was comprised of a total of 105 samples. The result of these nasopharyngeal specimens was already established by Reverse-Transcriptase Polymerase Chain Reaction (RT-PCR). Then the same (nasopharyngeal) specimens were analysed on Rapid Antigen Detection Test Device.
 Results: Out of 105 specimens, we got 88(83%) PCR positive samples and 17(16%) PCR negative, which, when compared with the Abbott PanbioTM Rapid Antigen Test Device, showed 60(57%) positive and 45(42.8%) negative samples having a sensitivity of 75(86%) (CI 67.04%-83.32%) and specificity of 100%(80.49-100%). The accuracy of the test was found to be 78(95%). This assay was found convenient and more appropriate for outdoor settings.Keywords: COVID-19, RT-PCR, Rapid antigen test, Specificity, Sensitivity.

Could oral hygiene prevent cases of at-home-acquired Legionnaires' disease? - Results of a comprehensive case-control study on infection sources, risk, and protective behaviors.

The "LeTriWa study" on community-acquired cases of Legionnaires' disease (LD) found that most cases likely acquired their infection at home (AHALD). However, which sources confer the infection is largely unknown. We therefore analyzed the data set from the LeTriWa study to find out if individual sources were associated with AHALD and if specific behavioral habits may increase or lower the risk for AHALD. During the study we had used two comparison groups: (i) controls matched for age group and hospital ("controls"), (ii) household members of cases with AHALD ("AHALD-HHM"). We inquired about exposure to water sources, such as showering or wearing dentures, as well as behavioral factors and habits related to oral hygiene. We took standardized household bathroom water and biofilm samples of both cases with AHALD and controls, and in addition from households of cases with AHALD only samples from suspect residential (non-)drinking water sources. We first conducted bivariate analyses for infection sources and behaviors, followed by multivariable analyses. There were 124 cases with AHALD, 217 controls and 59 AHALD-HHM. In bivariate analyses using controls for comparison, wearing dentures was the only variable significantly positively associated (odds ratio (OR) = 1.7, 95% confidence interval (CI) = 1.1-2.7, p-value = 0.02). Behavioral factors such as showering, letting water run before use and not being alcohol abstinent were significantly negatively associated, smoking was significantly positively associated. In a multivariable analysis, we identified good oral hygiene as a preventive factor for both denture wearers (OR = 0.33, 95% CI = 0.13-0.83, p-value = 0.02) and non-denture wearers (OR = 0.32, 95% CI = 0.10-1.04, p-value = 0.06). Analyses of comparisons with AHALD-HHM showed similar effects but lacked statistical power. We identified Legionella in 16 residential (non-)drinking water sources, one of which was a PCR-positive scratch sample of dentures. Wearing (inadequately cleaned) dentures or poor oral hygiene might confer an increased risk for AHALD, and oral hygiene may prevent AHALD. The hypothesis that Legionella in oral biofilm or dental plaque may be the cause of cases with AHALD should be examined further. If confirmed this may open new and simple avenues for the prevention of LD.

Single-Point Nail Sampling to Diagnose Onychomycosis Caused by Non-Dermatophyte Molds: Utility of Polymerase Chain Reaction (PCR) and Histopathology

The three most commonly used methods for diagnosing non-dermatophyte mold (NDM) onychomycosis are culture, polymerase chain reaction (PCR), and histopathology. Toenail samples from 512 patients (1 sample/patient) with suspected onychomycosis were examined using all three diagnostic tests. A statistically significant association was found between PCR and histopathology results, as well as between fungal culture and histopathology results. All PCR-positive and culture-positive dermatophyte samples were confirmed by histopathology. However, 15/116 (12.9%) of culture-positive NDM samples had negative histopathology results, while all PCR-positive NDM samples were confirmed by histopathology. The overall rate of dermatophyte detection was higher using PCR compared to culture (38.9% vs. 11.7%); the lower rate of NDM detection by PCR (11.7% vs. 38.9%) could be attributed to the restriction of the assay design to seven pre-selected targets. When repeat sampling in the clinic is not possible, a combination of NDM detection by PCR and positive histopathology of hyphae may be a proxy for NDM infection, particularly where the NDM occurs without a concomitant dermatophyte. There was a high degree of correlation between negative PCR and negative histopathology. A negative PCR result with negative histopathology findings may be a reliable proxy for the diagnosis of non-fungal dystrophy.

Dengue virus infection during window period of consecutive outbreaks in Nepal and assessment of clinical parameters

Nepal is an endemic country for dengue infection with rolling of every 3 year’s clear cyclic outbreaks with exponential growth since 2019 outbreak and the virus gearing towards the non-foci temperate hill regions. However, the information regarding circulating serotype and genotype is not frequent. This research discusses on the clinical features, diagnosis, epidemiology, circulating serotype and genotype among 61 dengue suspected cases from different hospitals of Nepal during the window period 2017–2018 between the two outbreaks of 2016 and 2019. E-gene sequences from PCR positive samples were subjected to phylogenetic analysis under time to most recent common ancestor tree using Markov Chain Monte Carlo (MCMC) and BEAST v2.5.1. Both evolution and genotypes were determined based on the phylogenetic tree. Serotyping by Real-time PCR and Nested PCR showed the co-circulation of all the 3 serotypes of dengue in the year 2017 and only DENV-2 in 2018. Genotype V for DENV-1 and Cosmopolitan Genotype IVa for DENV-2 were detected. The detected Genotype V of DENV-1 in Terai was found close to Indian genotype while Cosmopolitan IVa of DENV-2 found spreading to geographically safe hilly region (now gripped to 9 districts) was close to South-East Asia. The genetic drift of DENV-2 is probably due to climate change and rapid viral evolution which could be a representative model for high altitude shift of the infection. Further, the increased primary infection indicates dengue venturing to new populations. Platelets count together with Aspartate transaminase and Aalanine transaminase could serve as important clinical markers to support clinical diagnosis. The study will support future dengue virology and epidemiology in Nepal.

Investigation of the environmental presence of Salmonella spp. in finishing pigs at commercial swine farms in Kansas(USA).

Salmonellosis remains a major foodborne disease threat to public health worldwide. Swine are considered a reservoir for many Salmonella serotypes affecting humans; however, not all serotypes of concern in food animal products cause clinical signs of infection in swine. The objective of this study was to evaluate the presence and distribution of Salmonella spp. in finishing pigs at commercial farms across Kansas (USA). Five farms were selected and sampled when pigs weighed between 125 and 136 kg. Samples were collected and transported to the laboratory for processing following USDA-FSIS guidelines. Susceptibility and resistance profiles were also studied. Fifty-three percent (100/186) of samples were culture positive for Enterobacteriaceae, and 14% (14/100) were confirmed Salmonella positive by PCR with three of five farms having no PCR-positive samples. Salmonella serotype Braenderup was the most common serovar identified in environmental samples, while Salm. Infantis, Agona, and Montevideo were identified in fecal samples. Multidrug resistance patterns were only found in Farm 3, in fecal samples and in one floor sample. The observations reported in this study highlight areas of concern, such as locations prone to fecal contamination, to be considered when cleaning and sanitizing between groups of pigs to decrease presence of Salmonella spp. in farm environments.

No Isolate, No Problem: Using a Novel Insertion Sequence PCR to Link Rats to Human Shigellosis Cases in an Underserved Urban Community.

During an investigation into a cluster of Shigella flexneri serotype 2a cases in an underserved community, we assessed the relatedness of human and rat S. flexneri isolates utilizing a novel PCR targeting insertion sites (IS-PCR) of mobile elements in the Shigella genome characteristic of the cluster strain. Whole-genome sequences of S. flexneri (n = 50) associated with the cluster were analyzed. De novo genome assemblies were analyzed by a Geneious V10.2.6 motif search, and two unique IS were identified in all human Shigella sequences of the local cluster. Hydrolysis probe PCR assays were designed to detect these sequences consisting of forward and reverse primers to amplify across each insertion site and a hydrolysis probe spanning the insertion site. IS-PCR was performed for three Shigella PCR-positive culture-negative rat intestine specimens from this community. Both insertion sites were detected in the de novo genome assemblies of all clinical S. flexneri isolates (n = 50). Two of the three PCR-positive culture-negative rat samples were positive for both unique ISs identified in the human S. flexneri isolates, suggesting that the rat Shigella species strains were closely related to the human strains in the cluster. The cycle threshold (Ct) values were >35, indicating that the bacterial load was very low in the rat samples. Two unique IS were identified in clinical isolates from a community S. flexneri cluster. Both IS targets were identified in PCR-positive (Shigella spp.), culture-negative rat tissue and clinical isolates from humans, indicating relatedness. IMPORTANCE This article describes a novel molecular method to show relatedness between bacterial infections, which may not be able to grow in the laboratory due to treatment with antibiotics or for bacteria requiring unique conditions to grow well. Uniquely, we applied this technique to Shigella isolates from human cases associated with a local cluster in an underserved community, as well as rat samples from the same community. We believe that this novel approach can serve as a complementary method to support outbreak/cluster investigation for Shigella spp.

Impact of nanopore-based metagenome sequencing on tick-borne virus detection.

We evaluated metagenomic nanopore sequencing (NS) in field-collected ticks and compared findings from amplification-based assays. Forty tick pools collected in Anatolia, Turkey and screened by broad-range or nested polymerase chain reaction (PCR) for Crimean-Congo Hemorrhagic Fever Virus (CCHFV) and Jingmen tick virus (JMTV) were subjected to NS using a standard, cDNA-based metagenome approach. Eleven viruses from seven genera/species were identified. Miviruses Bole tick virus 3 and Xinjiang mivirus 1 were detected in 82.5 and 2.5% of the pools, respectively. Tick phleboviruses were present in 60% of the pools, with four distinct viral variants. JMTV was identified in 60% of the pools, where only 22.5% were PCR-positive. CCHFV sequences characterized as Aigai virus were detected in 50%, where only 15% were detected by PCR. NS produced a statistically significant increase in detection of these viruses. No correlation of total virus, specific virus, or targeted segment read counts was observed between PCR-positive and PCR-negative samples. NS further enabled the initial description of Quaranjavirus sequences in ticks, where human and avian pathogenicity of particular isolates had been previously documented. NS was observed to surpass broad-range and nested amplification in detection and to generate sufficient genome-wide data for investigating virus diversity. It can be employed for monitoring pathogens in tick vectors or human/animal clinical samples in hot-spot regions for examining zoonotic spillover.

Identification and genotyping of Acanthamoeba spp. in the water resources of western Iran

BackgroundAcanthamoeba spp. is opportunistic amoeba that resides in water, soil, and air. Some pathogenic genotypes of the genus of Acanthamoeba can cause granulomatous amoebic encephalitis (GAE) in people with a defective immune system. The parasite can also cause Acanthamoeba keratitis (AK) among contact lens users. This study was conducted to isolate and identify the Acanthamoeba genotypes in water resources in Lorestan province, western Iran. MethodsCollected 72 water samples from surface and groundwater (springs and aqueducts) in Lorestan province. Samples were filtered and cultured in non-nutrient 1.5% agar medium covered with Escherichia coli (E. coli) at 25 °C. DNA extraction was done and the PCR reaction was performed to detect the Acanthamoeba spp. The positive PCR products were sequenced to determine the genotypes of Acanthamoeba. ResultsOut of 72 examined water samples, 23.61% were positive for Acanthamoeba sp. by PCR. From PCR-positive samples, 8 (47.05%) samples were T4 genotypes and others were other Acanthamoeba genotypes (T1-T23). Therefore, approximately half of the genotypes belong to the pathogenic T4 genotype. ConclusionsThe water examined samples in western provinces of Iran have the potential risk factor for public health. Therefore, the efforts of healthcare providers are needed to identify, train, and prevention from human infections.

Molecular characterisation of the current high prevalence of the new CPV-2c variants in the Southern Vietnamese dogs signifies a widespread in the worldwide dog population.

Canine parvovirus type 2 (CPV-2) is known as the primary etiological agent cause of acute gastroenteritis, myocarditis and death of canids worldwide. In Vietnam, although CPV-2 infection and its outbreaks are the most important risk factors of the canine's health concern, lack of available information about the molecular epidemiology of the CPV-2. In this study, the complete coding sequences of 10 CPV-2 strains collected from dogs vaccinated with CPV-2 vaccination were analysed to better understand the genomic characteristics of the current circulating CPV-2 in Vietnam. Ten CPV-specific PCR-positive rectal swab samples were collected from dogs with acute symptoms of haemorrhagic diarrhoea and vomiting in Vietnam in 2019. The complete coding sequences of these CPV strains were analysed to determine their phylogeny and genetic relationship with other available CPV strains globally. Analysis of the VP2 gene sequences demonstrated that the studied strains belonged to the new CPV-2c variants with the unique mutations at amino acids 5Ala-Gly and 447Iso-Met . Phylogenetic tree analysis indicated that the studied strains share a common evolutionary origin with the current CPV-2c strains circulating in dogs in Asia countries, including China, Thailand, Taiwan and Mongolia, in recent years. Low sequence identity between the studied strains and commercial vaccine strains was observed. This study provides deep insights into the molecular characteristics, genetic diversity, and evolution of circulating CPV-2 strains in Vietnam. We recommend more studies to estimate the effectiveness of the CPV vaccine and the need to continue developing other effective vaccination essential to better control the widespread of these new CPV-2 variants.

Post-exposure prophylaxis with SA58 (anti-SARS-COV-2 monoclonal antibody) nasal spray for the prevention of symptomatic COVID-19 in healthy adult workers: a randomized, single-blind, placebo-controlled clinical study*

ABSTRACT Monoclonal antibodies (mAbs) and the post-exposure prophylaxis (PEP) with mAbs represent a very important public health strategy against coronavirus disease 2019 (COVID-19). This study has assessed a new Anti-SARS-COV-2 mAb (SA58) Nasal Spray for PEP against COVID-19 in healthy adults aged 18 years and older within three days of exposure to a SARS-CoV-2 infected individual. Recruited participants were randomized in a ratio of 3:1 to receive SA58 or placebo. Primary endpoints were laboratory-confirmed symptomatic COVID-19 within the study period. A total of 1222 participants were randomized and dosed (SA58, n = 901; placebo, n = 321). Median of follow-up was 2.25 and 2.79 days for SA58 and placebo, respectively. Adverse events occurred in 221 of 901 (25%) and 72 of 321 (22%) participants with SA58 and placebo, respectively. All adverse events were mild in severity. Laboratory-confirmed symptomatic COVID-19 developed in 7 of 824 participants (0.22 per 100 person-days) in the SA58 group vs. 14 of 299 (1.17 per 100 person-days) in the placebo group, resulting in an estimated efficacy of 80.82% (95%CI 52.41%−92.27%). There were 32 SARS-CoV-2 reverse transcriptase polymerase chain reaction (RT–PCR) positives (1.04 per 100 person-days) in the SA58 group vs. 32 (2.80 per 100 person-days) in the placebo group, resulting in an estimated efficacy of 61.83% (95%CI 37.50%−76.69%). A total of 21 RT–PCR positive samples were sequenced and all were the Omicron variant BF.7. In conclusion, SA58 Nasal Spray showed favourable efficacy and safety in preventing symptomatic COVID-19 or SARS-CoV-2 infection in adults who had exposure to SARS-CoV-2 within 72 h.

Foodborne protozoan parasites in fresh mussels and oysters purchased at retail in Canada

Studies worldwide have reported the presence of protozoan parasites in a variety of commercial bivalve shellfish. The uptake of these parasites by shellfish occurs during filter feeding in faecally-contaminated waters. The objective of the present study was to determine the prevalence of Giardia, Cryptosporidium and Toxoplasma in fresh, live shellfish purchased in three Canadian provinces as part of the retail surveillance activities led by FoodNet Canada (Public Health Agency of Canada). Packages containing mussels (n = 253) or oysters (n = 130) were purchased at grocery stores in FoodNet Canada sentinel sites on a biweekly basis throughout 2018 and 2019, and shipped in coolers to Health Canada for testing. A small number of packages were not tested due to insufficient quantity or poor quality. Following DNA extraction from homogenized, pooled tissues, nested PCR and DNA sequencing were used to detect parasite-specific sequences. Epifluorescence microscopy was used to confirm the presence of intact cysts and oocysts in sequence-confirmed PCR-positive samples. Giardia duodenalis DNA was present in 2.4 % of 247 packages of mussels and 4.0 % of 125 packages of oysters, while Cryptosporidium parvum DNA was present in 5.3 % of 247 packages of mussels and 7.2 % of 125 packages of oysters. Toxoplasma gondii DNA was only found in mussels in 2018 (1.6 % of 249 packages). Parasite DNA was detected in shellfish purchased in all three Canadian provinces sampled, and there was no apparent seasonal variation in prevalence. While the present study did not test for viability, parasites are known to survive for long periods in the marine environment, and these findings suggest that there is a risk of infection, especially when shellfish are consumed raw.

Molecular investigation of malaria-infected patients in Djibouti city (2018–2021)

BackgroundThe Republic of Djibouti is a malaria endemic country that was in pre-elimination phase in 2006–2012. From 2013, however, malaria has re-emerged in the country, and its prevalence has been increasing every year. Given the co-circulation of several infectious agents in the country, the assessment of malaria infection based on microscopy or histidine-rich protein 2 (HRP2)-based rapid diagnostic tests (RDT) has shown its limitations. This study, therefore, aimed to assess the prevalence of malaria among febrile patients in Djibouti city using more robust molecular tools.MethodsAll suspected malaria cases reported to be microscopy-positive were randomly sampled (n = 1113) and included in four health structures in Djibouti city over a 4-year period (2018–2021), mainly during the malaria transmission season (January–May). Socio-demographic information was collected, and RDT was performed in most of the included patients. The diagnosis was confirmed by species-specific nested polymerase chain reaction (PCR). Data were analysed using Fisher’s exact test and kappa statistics.ResultsIn total, 1113 patients with suspected malaria and available blood samples were included. PCR confirmed that 788/1113 (70.8%) were positive for malaria. Among PCR-positive samples, 656 (83.2%) were due to Plasmodium falciparum, 88 (11.2%) Plasmodium vivax, and 44 (5.6%) P. falciparum/P. vivax mixed infections. In 2020, P. falciparum infections were confirmed by PCR in 50% (144/288) of negative RDTs. After the change of RDT in 2021, this percentage decreased to 17%. False negative RDT results were found more frequently (P < 0.05) in four districts of Djibouti city (Balbala, Quartier 7, Quartier 6, and Arhiba). Malaria occurred less frequently in regular bed net users than in non-users (odds ratio [OR]: 0.62, 95% confidence interval [CI]: 0.42–0.92).ConclusionsThe present study confirmed the high prevalence of falciparum malaria and, to a lesser extent, vivax malaria. Nevertheless, 29% of suspected malaria cases were misdiagnosed by microscopy and/or RDT. There is a need to strengthen the capacity for diagnosis by microscopy and to evaluate the possible role of P. falciparum hrp2 gene deletion, which leads to false negative cases of P. falciparum.