WITHDRAWN: Overview of dengue virus infection in Saudi Arabia

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Insights into gastrointestinal virome : etiology and public exposure

Funding: The authors extend their appreciation to the Deanship of Scientific Research at King Saud University for funding this work through research group no. RG-1441-492.Recycled wastewater is widely used owing to the potential shortage of water resources for drinking purposes, recreational activities, and irrigation. However, gut microbiomes of both human beings and animals negatively affect this water quality. Wastewater contamination is continuously monitored, using fecal contamination indicators or microbial source tracking approaches, to oppose arising enteric infections. Viral gastroenteritis is considered a principal manifestation of waterborne pathogenic virome-mediated infections, which are mainly transmitted via the fecal-oral route. Furthermore, acquired enteric viromes are the common cause of infantile acute diarrhea. Moreover, public exposure to wastewater via wastewater discharge or treated wastewater reuse has led to a significant surge of public health concerns. In this review, we discussed the etiology of waterborne enteric viromes, notably gastrointestinal virus infections, and public exposure to municipal wastewater. Conclusively, the early human virome is affected mainly by birth mode, dietary behavior, and maternal health, and could provide a signature of disease incidence, however, more virome diversification is acquired in adulthood. A multi-phase treatment approach offered an effective means for the elimination of wastewater reuse mediated public risks. The insights highlighted in this paper offer essential information for defining probable etiologies and assessing risks related to exposure to discharged or reused wastewater.Publisher PDFPeer reviewe

Non-structural proteins of arthropod-borne bunyaviruses: roles and functions

Viruses within the Bunyaviridae family are tri-segmented, negative-stranded RNA viruses. The family includes several emerging and re-emerging viruses of humans, animals and plants, such as Rift Valley fever virus, Crimean-Congo hemorrhagic fever virus, La Crosse virus, Schmallenberg virus and tomato spotted wilt virus. Many bunyaviruses are arthropod-borne, so-called arboviruses. Depending on the genus, bunyaviruses encode, in addition to the RNA-dependent RNA polymerase and the different structural proteins, one or several non-structural proteins. These non-structural proteins are not always essential for virus growth and replication but can play an important role in viral pathogenesis through their interaction with the host innate immune system. In this review, we will summarize current knowledge and understanding of insect-borne bunyavirus non-structural protein function(s) in vertebrate, plant and arthropod

Isolation of Thermoalkalophilic-?-amylase Producing Bacteria and Optimization of Potato Waste Water Medium for Enhancement of ?-amylase Production

Sixty one thermoalkalophilic bacteria were isolated from soil samples in Saudi Arabia’s southern region. Isolate TA-38, obtained from the Tanomah region, showed the best performance for enzyme production and was submitted for further study. It was identified as Bacillus axarquiensis based on 16S rRNA gene sequencing studies. The feasibility of using potato waste water as a simple and cheap medium for the production of ?-amylase was evaluated compared with starch broth medium. The production of ?-amylase in the potato waste water medium was only 13.8% less than that of the starch medium. Maximum enzyme production was achieved after 48 hours of cultivation at the beginning of the stationary phase at pH 10.0 and 50 0C. The appropriate addition of starch; nitrogen; phosphate; and calcium to potato waste water significantly enhanced the production of ?-amylase. The enzyme production reached a maximum of 64.5 Uml-1 with the potato wastewater adding with 0.5 % starch; 0.4 % yeast extract; 0.04% CaCl2-2H2O and 0.05 % KH2PO4.  The optimization of the potato waste water medium led to an approximately 4.02 fold increase in the production of ?-amylase compared to starch broth medium. Data indicated that the potato waste water contained substrates which could be used by bacterial isolate for the production of ?-amylase production and the developed procedure was cost effective since it requires only a slightly addition of nutrients to the medium. Keywords: Isolation; ?-amylase; 16S rRNA; Production; Potato waste water; Thermoalkaliphilic bacteria

Distribution and molecular identification of Culex pipiens and Culex tritaeniorhynchus as potential vectors of Rift Valley fever virus in Jazan, Saudi Arabia

Entomologic investigations were conducted in the Al-Darb, Al-Reath, Al-Aridah, Abuareesh, Al-Ahad, Samttah, Sabyah, Damad and Beash areas by CO2-baited CDC miniature light traps in the Jazan region. Vectors were identified morphologically, as well as COI gene segment amplification and sequencing. The relative abundance (RA%) and pattern of occurrence (C%) were recorded. The presence of the Rift Valley fever virus (RVFV) in pooled mosquito samples was investigated by reverse transcriptase-polymerase chain reaction (RT-PCR). Culex pipiens (C. pipiens) and Culex tritaeniorhynchus (C. tritaeniorhynchus) were found with RA% values of 96% and 4%, respectively, in the region. Significant variations in vector population densities were observed in different districts. The C. pipiens was found highly abundant in all districts and RA% value (100%) was recorded in the Al-Darb, Al-Reath, Al-Aridah, Samttah and Damad areas, whereas RA% values (93.75%, 93.33%, 92.30% and 91.66%) were noted in Al-Ahad, Sabyah, Abuareesh and Beash districts, respectively. RA% values for C. tritaeniorhynchus were recorded as 8.33%, 7.70%, 6.66% and 6.25% in Beash, Abuareesh, Sabyah and Al-Ahad areas, respectively. The pattern of occurrence for C. pipiens and C. tritaeniorhynchus was recorded as 100% and 44.4% in the region. Phylogenetic analysis of C. pipiens and C. tritaeniorhynchus exhibited a close relationship with mosquitoes from Kenya and Turkey, respectively. All mosquito samples tested by RT-PCR were found negative for RVFV. In summary, the current study assessed the composition, abundance, distribution of different mosquito vectors and presence of RVFV in different areas of the Jazan region. Our data will help risk assessments of RVFV future re-emergence in the region

Functional analysis of the orthobunyavirus nucleocapsid (N) protein

Bunyamwera virus (BUNV) is the prototype of the family Bunyaviridae. It has a tripartite genome consisting of negative sense RNA segments called large (L), medium (M) and small (S). The S segment encodes the nucleocapsid protein (N) of 233 amino acids. The N protein encapsidates all three segments to form transcriptionally active ribonucleoproteins (RNPs). The aim of this project was to determine the domain map of BUNV N protein. To investigate residues in BUNV N crucial for its functionality, random and site- specific mutagenesis were performed on a cDNA clone encoding the BUNV N protein. In total, 102 single amino acid substitutions were generated in the BUNV N protein sequence. All mutant N proteins were used in a BUNV minigenome system to compare their activity to wt BUNV N. The mutant proteins displayed a wide-range of activity, from parental-like to essentially inactive. The most disruptive mutations were R94A, I118N, W134A, Y141C, L177A, K179I and W193A. Sixty-four clones carrying single substitutions in the BUNV N protein were used in the BUNV rescue system in an attempt to recover viable mutant viruses. Fifty recombinant mutant viruses were rescued and 14 N genes were nonrescuable. The 50 mutant viruses were characterized by: titration, protein labelling, western blotting, temperature sensitivity and host-restriction. Mutant viruses displayed a wide range of titers between 10³ -10⁸ pfu/ml, and three different plaque sizes large, medium and small. Protein labelling and western blotting showed that mutations in the N gene did not affect expression of the other viral genes as much as affecting N protein expression. It was demonstrated that single amino acid substitutions could alter N protein electrophoretic mobility in SDS- PAGE (e.g. P19Q and L53F). Temperature sensitivity tests showed that recombinant viruses N74S, S96S, K228T and G230R were ts, growing at 33˚C but not at 37˚C or 38˚C, while the parental virus grew at all temperatures. Using the northern blotting technique, mutant viruses N74S and S96G were shown to have a ts defect in genome-synthesis (late replication step), while mutant viruses K228T and G230R had a ts defect in antigenome- synthesis (early replication step). Host-restriction experiments were performed using 5 different cell lines (Vero-E6, BHK-21, 2FTGH-V, A549-V and 293-V). Overall, the parental virus grew similarly in all cell lines. Likewise, the majority of mutant viruses follow this pattern except mutant virus Y23A. It showed a 100-fold reduction in titer in 2FTGH-V cells. Comparing the ratios of intracellular and extracellular particles revealed that only 15% of the total virus particles of mutant Y23A was released as extracellular particles compared to 30% of the parental virus. Fourteen N genes were nonrescuable. They were characterized by (i) their activity in the BUNV minigenome system, (ii) their activity in BUNV packaging assay, (iii) their ability to form multimers, (iv) their ability to interact with L protein, and (v) their impact on RNA synthesis. In summary, BUNV N protein was shown to be multi-functional and involved in the regulation of virus transcription and replication, RNA synthesis and assembly, via interactions with the viral L polymerase, RNA backbone, itself or the viral glycoproteins

Non-Structural Proteins of Arthropod-Borne Bunyaviruses: Roles and Functions

Viruses within the Bunyaviridae family are tri-segmented, negative-stranded RNA viruses. The family includes several emerging and re-emerging viruses of humans, animals and plants, such as Rift Valley fever virus, Crimean-Congo hemorrhagic fever virus, La Crosse virus, Schmallenberg virus and tomato spotted wilt virus. Many bunyaviruses are arthropod-borne, so-called arboviruses. Depending on the genus, bunyaviruses encode, in addition to the RNA-dependent RNA polymerase and the different structural proteins, one or several non-structural proteins. These non-structural proteins are not always essential for virus growth and replication but can play an important role in viral pathogenesis through their interaction with the host innate immune system. In this review, we will summarize current knowledge and understanding of insect-borne bunyavirus non-structural protein function(s) in vertebrate, plant and arthropod

Mutational Analysis of the Bunyamwera Orthobunyavirus Nucleocapsid Protein Gene▿ †

The bunyavirus nucleocapsid protein, N, is a multifunctional protein that encapsidates each of the three negative-sense genome segments to form ribonucleoprotein complexes that are the functional templates for viral transcription and replication. In addition, N protein molecules interact with themselves to form oligomers, with the viral L (RNA polymerase) protein, with the carboxy-terminal regions of either or both of the virion glycoproteins, and probably also with host cell proteins. Bunyamwera virus (BUNV), the prototype bunyavirus, encodes an N protein of 233 amino acids in length. To learn more about the roles of individual amino acids in the different interactions of N, we performed a wide-scale mutagenic analysis of the protein, and 110 single-point mutants were obtained. When the mutants were employed in a minireplicon assay to examine their effects on viral RNA synthesis, a wide range of activities compared to those of wild-type N protein were observed; changes at nine amino acid positions resulted in severely impaired RNA synthesis. Seventy-seven mutant clones were selected for use in the bunyavirus reverse genetics system, and 57 viable recombinant viruses were recovered. The recombinant viruses displayed a range of plaque sizes and titers in cell culture (from approximately 103 to 108 PFU/ml), and a number of viruses were shown to be temperature sensitive. Different assays were applied to determine why 20 mutant N proteins could not be recovered into infectious virus. Based on these results, a preliminary domain map of the BUNV N protein is proposed