Seroprevalence of Toxoplasma gondii infection in feral cats in Qatar

Background: Cats are essential in the life cycle of Toxoplasma gondii as they can shed the environmentally resistant oocysts after acquiring infection. Human populations living in cities with high densities of feral cats are therefore likely to be at risk of infection. The current study is the first to estimate the seroprevalence of T. gondii in the feral cat population in Qatar. We investigated the seroprevalence of T. gondii among 495 adult cats from urban and suburban districts in Qatar. Using results from the Modified Agglutination Test, we fitted statistical models with host sex, area and season as explanatory factors and seropositivity as the outcome. Results: The analysis revealed an overall seroprevalence of 82%. Seroprevalence was significantly higher in the summer season (P = 0.006). No significant difference was detected (P > 0.05) between seroprevalence in female and male cats and in cats from urban and suburban districts of Qatar. Conclusions: Despite the seasonal difference, the observed seroprevalence of T. gondii suggests high environmental contamination throughout the year, with some female cats generating more intense responses compared to males. Both findings merit further investigations.NPRP grant number NPRP 4-164-4-001 from Qatar National Research Fun

Cryptosporidium spp., prevalence, molecular characterisation and socio-demographic risk factors among immigrants in Qatar.

The World Health Organization WHO has estimated that in developed countries, up to 30% of the population may suffer from foodborne diseases each year, and that in developing countries up to 2 million deaths per annum can be attributed to cryptosporidiosis. Reports have already emphasized the role of immigrants in outbreaks of parasitic diseases especially those working in food processing industries. Herein we assessed Cryptosporidium spp. infections among immigrants in Qatar with a special focus on food handlers and housemaids. The overall prevalence of Cryptosporidium spp. by q-PCR among 839 asymptomatic subjects was 4.5%. Based on the Gp60 gene, the majority of isolates were identified as C. parvum subtype IIdA20G1b. The positive sample for C. hominis was subtyped as IeA12G3T3. Seven mixed infections were also identified (four C. parvum + C. hominis, and three C. parvum + C. meleagridis). The prevalence of Cryptosporidium spp. did not differ significantly between the sexes or age classes but varied significantly between subjects affiliated to different religions with the lowest prevalence among the Muslims. Multifactorial analysis retained also marked significance with education, income, and a house contents index. Our results contribute to a better understanding of the epidemiology of cryptosporidiosis and the risk factors associated with the likelihood of carrying this infection among immigrant workers from developing countries.QNR

Molecular Analysis of Aquaglyceroporin 1 Gene in Non-Healing Clinical Isolates Obtained from Patients with Cutaneous Leishmaniasis from Central of Iran

Background: Regarding the antimonial-resistant of Leishmania spp., understanding of related mechanism is necessary. One of the most important involved molecules is aquaglyceropin1 (AQP1). The aim of this study was molecular analysis of AQP1 gene from antimonial-resistant clinical isolates and its expression. Methods: Overall, 150 patients with cutaneous leishmaniasis referring to the reference laboratories of Yazd and Varzaneh,, located 105km southeast of Isfahan and 240km away from Yazd, were assessed from Jun 2015 to Dec 2017. After sampling, staining was done and evaluated for Leishman by microscope. Samples were collected in RNAlater solution for gene expression analysis in non-healing isolates. DNA extraction was performed from each slide with Leishman body. All patients with L. major isolates detected by ITS1-PCR-RFLP were followed for finding the resistant isolates, consequence of molecular characterization of AQP1 using PCR-RFLP. Gene expression of AQP1 from all resistant isolates was assessed in comparison with the one in a sensitive isolate. Statistical analysis was done using SPSS. The significance level was considered ≤0.05. Results: Five isolates were detected as antimonial resistant. Molecular detection and identification were appeared that all were L. major. The molecular characterization of AQP1 showed G562A mutation. Gene expression of AQP1 in resistant isolates showed 1.67 fold higher than the sensitive isolate. Conclusion: We reported a new point mutation of G562A in AQP1 gene involved in molecular mechanism in resistant isolates

Tunisian Toxoplasma gondii strains genotyping by the use of AK69 marker

<p>Abstract</p> <p>Background</p> <p>Clinical manifestation due to infection by <it>Toxoplasma gondii </it>is closely linked to the infecting strain of the parasite. Several genetic markers are available to determinate its genotype but few of them are able to discriminate between the three predominant lineages, namely types I, II and III. The number of markers decreases when atypical, recombinant/mixed genotypes need to be identified.</p> <p>Findings</p> <p>In our study, the contribution of sequence polymorphisms in the AK69 gene as typing markers for <it>T. gondii </it>was investigated for the first time in an epidemiological study. The coding region of the marker was amplified, sequenced and aligned for different <it>Toxoplasma </it>strains. The identified nucleotide polymorphism at 12 positions was able to highly discriminate between the different congenital toxoplasmosis Tunisian strains. Moreover the high detection sensitivity level of the marker enabled unambiguous identification of mixed/recombinant genotypes directly.</p> <p>Conclusion</p> <p>It can be, thus, very useful for direct typing in areas where such genotypes are frequently encountered, mainly in the African continent.</p

Helminth infections among long-term residents and settled immigrants in Qatar in the decade from 2005 to 2014: temporal trends and varying prevalence among subjects from different regional origins

Background: Travel and migration from developing regions, where tropical diseases are common, to more developed industrialised nations can contribute to the introduction and subsequent spread of infections. With its rapidly expanding economy, Qatar has attracted vast numbers of immigrant workers in the last two decades, often from countries with poor socio-economic levels. Many used to arrive with patent intestinal parasitic infections. Methods: We analysed the prevalence of helminth infections in a dataset of 29,286 records of subjects referred for stool examination at the Hamad Medical Corporation over the course of a decade (2005 to 2014, inclusive). Results: Overall prevalence of combined helminth infections was low (1.86 %) but there were significant temporal trends, age and sex effects and those arising from the region of origin of the subjects. The most common helminths were hookworms (overall prevalence 1.22 %), which accounted for 70.1 % of cases, and therefore patterns for combined helminth infections were largely driven by hookworms. In both cases, and also in Trichuris trichiura and Ascaris lumbricoides, prevalence peaked in 2008, since when prevalence has been steadily falling. Helminth infections were largely concentrated among subjects from five Asian countries (Nepal, Bangladesh, Sri Lanka, India and Pakistan), and there was a highly biased prevalence in favour of male subjects in all cases. Prevalence of all three nematodes peaked in age class 7 (mean age 25.5 years, range = 20–29) and there were significant interactions between region of origin, sex of subjects and prevalence of hookworms. Conclusion: These results offer optimism that prevalence will continue to decline in the years ahead, especially if control is targeted at those most at risk of carrying infections.Qatar National Research Fund (QRNF) at Qatar Foundation for supporting this study through the National Priorities Research Program (NPRP) (Project No. NPRP 4-1283-3-327)

Evaluation of reverse transcription-loop-mediated isothermal amplification (RT-LAMP) for detecting SARS-CoV-2 in clinical, environmental and animal samples.

Background: First described 20 years ago by Notomi et al., the loop-mediated isothermal amplification (LAMP) assay is robust, rapid and straightforward, yet retains high sensitivity and specificity. These features have seen the LAMP assay and the inclusion of a reverse transcriptase (RT-LAMP) implemented for a broad range of molecular diagnostic applications extending from infectious diseases, including detection of the original SARS-CoV-1 virus. The advantages of RT-LAMP include using different reagents than RT-qPCR, the potential for direct processing of samples without the need for prior RNA extraction and an extremely rapid turn-around time. Several groups have now described different RT-LAMP assays for detection of SARS-CoV-2 RNA. Therefore, the aim of this study is to assess the feasibility, sensitivity and effectiveness of LAMP technique in detecting SARS-CoV-2 in different type of samples. Method: New England Biolabs (NEB) LAMP master mixes were used. Six set of primers specific to SARS-CoV-2 were obtained from IDT. The reaction mix consisting of LAMP master mix, primer working solution and a sample was incubated at 65?C and results were collected after 30 mins. Results: In just 30 min we were able to detect the virus without any prior sample processing. Our primers were able to detect up to 100 copies of the viruses, which is comparable to the RT-PCR that we currently use in our lab. The primers were tested against all other coronavirus and they have shown 100% specificity to the novel SARS-CoV-2 virus. Both the florescent and calorimetric master mixes were able to detect the virus in all tested samples: clinical, animal and environmental. Conclusion: LAMP is a fast reliable technique that could be used as a quick screening method for the detection of SARS-CoV-2 in different settings and using different collection medium

Whole genome sequencing of marine organisms by Oxford Nanopore Technologies: Assessment and optimization of HMW-DNA extraction protocols

Marine habitats are Earth's largest aquatic ecosystems, yet little is known about marine organism's genomes. Molecular studies can unravel their genetics print, thus shedding light on specie's adaptation and speciation with precise authentication. However, extracting high molecular weight DNA from marine organisms and subsequent DNA library preparation for whole genome sequencing is challenging. The challenges can be explained by excessive metabolites secretion that co-precipitates with DNA and barricades their sequencing. In this work, we sought to resolve this issue by describing an optimized isolation method and comparing its performance with the most commonly reported protocols or commercial kits: SDS/phenol–chloroform method, Qiagen Genomic Tips kit, Qiagen DNeasy Plant mini kit, a modified protocol of Qiagen DNeasy Plant kit, Qiagen DNeasy Blood and Tissue kit, and Qiagen Qiamp DNA Stool mini kit. Our method proved to work significantly better for different marine species regardless of their shape, consistency, and sample preservation, improving Oxford Nanopore Technologies sequencing yield by 39 folds for Spirobranchus sp. and enabling generation of almost 10 GB data per flow cell/run for Chrysaora sp. and Palaemon sp. samples

Molecular investigation of waterborne protozoan contamination using marine Demospongiae

Sponges play important role within aquatic ecosystems due to their diverse abilities including filter-based feeding mechanisms. Hence, this study evaluated the potential use of sponges as ecological biomonitors for water safety surveillance, especially in the presence of Waterborne protozoan pathogens WBPP. Sponge specimens were collected from different Qatari marine ecosystems and subjected to gDNA extraction and real-time PCR using specific primer sets for the most common WBPP. Two sponges from the coastal marine ecosystems were found to be positive for Blastocystis sp., and one sponge was positive for Dientamoeba fragilis within offshore site. No Cryptosporidium spp., Giardia duodenalis, nor Toxoplasma gondii were detected. Further genotyping analysis revealed that the Blastocystis sp. positive samples were subtype ST3 (allele 34), which matched local clinical isolates and D. fragilis specimen was unambiguously clustering with Genotype 2. In conclusion, this study demonstrates the role of marine sponges as ecological biomonitors for WBPP screening and provide insights into these pathogens widespread and their potential transmission to marine and terrestrial organisms including human

Nanopore Sequencing SARS-CoV-2 genome in Qatar

Background: The current pandemic, COVID-19, is cause by an RNA coronavirus that was recently identified as SARS-CoV-2. RNA viruses tend to have a high mutation rate; the rate is around a million times greater than that of their hosts. The mutagenic potential of the virus depends on many factors, including the fidelity of nucleic acid-replicating viral enzymes, such as SARS-CoV-2 RNA dependent RNA polymerase (RdRp). The rate of mutation drives viral evolution and genome variability, consequently allowing viruses to escape the immunity of the host and develop resistance to drugs. Therefore, the characterization of SARS-CoV-2 variants might lead to implement better therapeutics treatments, vaccines design and identify new diagnostics approaches. Aim: The aim of this study was to establish a fast sequencing method to identify SARS-CoV-2 mutations in Qatar. This will help to assess if there are new viral variants that are spreading in country. Methods: RNA was isolated from samples collected from Qatar COVID-19 positive patients. The Artic Network V3 primer scheme and Oxford Nanopore ligation sequencing kit were used to prepare the sequencing libraries. Libraries were loaded on to R9.4.1 flow cells and ran on a GridION. Bioinformatics analysis was done following the Artic Network SARA-CoV-2 bioinformatics tools. Results: Genome coverage of sequenced samples was >80% and the depth was average at 200x. The coverage was highly dependable on sample viral load; samples of CT value lower than 30 resulted in better sequence coverage. The sequenced genomes were deposited in GISAID and were mainly clustering with genomes deposited from the UK. Sequences were compared to Illumina and sanger sequences and they showed compatible results. Conclusion: The use of ONT to sequence SARA-CoV-2 is a quick, affordable, and reliable technique to determine viral mutation. Using this technique, the first sequences from Qatar were deposited in to GISAID. Up to date, 700 genomes have been sequences from Qatari samples

Molecular analysis of the enteric protozoa associated with acute diarrhea in hospitalized children

Pediatric diarrhea is a common cause of death among children under 5 years of age. In the current study, we investigated the frequency of intestinal parasites among 580 pediatric patients with chronic diarrhea. Parasitic protozoa (all species combined) were detected by molecular tools in 22.9% of the children and the most common parasite was Cryptosporidiumspp. (15.1%). Blastocystis hominis was detected in 4.7%, Dientamoeba fragilis in 4%, Giardia duodenalis in 1.7%, and Entamoeba histolytica in 0.17%. Protozoan infections were observed among all regional groups, but prevalence was highest among Qatari subjects and during the winter season. Typing of Cryptosporidium spp. revealed a predominance of Cryptosporidium parvum in 92% of cases with mostly the IIdA20G1 subtype. Subtypes IIdA19G2, IIdA18G2, IIdA18G1, IIdA17G1, IIdA16G1, and IIdA14G1 were also detected. For Cryptosporidium hominis, IbA10G2 and IbA9G3 subtypes were identified. This study provides supplementary information for implementing prevention and control strategies to reduce the burden of these pediatric protozoan infections. Further analyses are required to better understand the local epidemiology and transmission of Cryptosporidium spp. in Qatar.This study was funded by the Qatar National Research Fund (QRNF) at Qatar Foundation through the National Priorities Research Program (Project No. NPRP8-1556-3-313)