Genomic characterization of SARS-CoV-2 in Egypt: insights into spike protein thermodynamic stability.

The overall pattern of the SARS-CoV-2 pandemic so far has been a series of waves; surges in new cases followed by declines. The appearance of novel mutations and variants underlie the rises in infections, making surveillance of SARS-CoV-2 mutations and prediction of variant evolution of utmost importance. In this study, we sequenced 320 SARS-CoV-2 viral genomes isolated from patients from the outpatient COVID-19 clinic in the Children's Cancer Hospital Egypt 57357 (CCHE 57357) and the Egypt Center for Research and Regenerative Medicine (ECRRM). The samples were collected between March and December 2021, covering the third and fourth waves of the pandemic. The third wave was found to be dominated by Nextclade 20D in our samples, with a small number of alpha variants. The delta variant was found to dominate the fourth wave samples, with the appearance of omicron variants late in 2021. Phylogenetic analysis reveals that the omicron variants are closest genetically to early pandemic variants. Mutation analysis shows SNPs, stop codon mutation gain, and deletion/insertion mutations, with distinct patterns of mutations governed by Nextclade or WHO variant. Finally, we observed a large number of highly correlated mutations, and some negatively correlated mutations, and identified a general inclination toward mutations that lead to enhanced thermodynamic stability of the spike protein. Overall, this study contributes genetic and phylogenetic data, as well as provides insights into SARS-CoV-2 viral evolution that may eventually help in the prediction of evolving mutations for better vaccine development and drug targets.

SARS-CoV-2 genome variations and evolution patterns in Egypt: a multi-center study.

A serious global public health emergency emerged late November 2019 in Wuhan City, China, by a new highly pathogenic virus, SARS-CoV-2. The virus evolution spread has been tracked by three developing databases: GISAID, Nextstrain and PANGO to understand its circulating variants. In this study, 110 diagnosed positive COVID-19 patient's samples, were collected from Kasr Al-Aini Hospital and the Children Cancer Hospital Egypt 57357 between May 2020 and January 2021, with clinical severity ranging from mild to severe. The viral genomes were sequenced by next generation sequencing, and phylogenetic analysis was performed to understand viral transmission dynamics. According to Nextstrain clades, most of our sequenced samples belonged to clades 20A and 20D, which in addition to clade 20B were present from the beginning of sample collection in May 2020. Clades 19A and 19B, on the other hand, appeared in the mid and late 2020 respectively, followed by the disappearance of clade 20B at the end of 2020. We identified a relatively high prevalence of the D614G spike protein variant and novel patterns of mutations associated together and with different clades. We also identified four mutations, spike H49Y, ORF3a H78Y, ORF8 E64stop and nucleocapsid E378V, associated with higher disease severity. Altogether, our study contributes genetic, phylogenetic, and clinical correlation data about the spread of the SARS-CoV-2 pandemic in Egypt.

Whole genome sequencing of Klebsiella pneumoniae clinical isolates sequence type 627 isolated from Egyptian patients.

Klebsiella pneumoniae is considered a threat to public health especially due to multidrug resistance emergence. It is largely oligoclonal based on multi-locus sequence typing (MLST); in Egypt, ST 627 was recently detected. Despites the global dissemination of this ST, there is still paucity of information about it. Herein, we used 4 K. pneumoniae ST627 for whole genome sequencing utilizing an Illumina MiSeq platform. Genome sequences were examined for resistance and virulence determinants, capsular types, plasmids, insertion sequences, phage regions, and Clustered Regularly Interspaced Palindromic Repeats (CRISPR) regions using bioinformatic analysis. The molecular characterization revealed 15 and 65 antimicrobial resistance and virulence genes, respectively. Resistance genes such as tet(D), aph(3'')-Ib, aph(6)-Id, blaTEM-234, fosA, and fosA6; were mainly responsible for tetracycline, aminoglycoside, and fosfomycin resistance; respectively. The capsular typing revealed that the four strains are KL-24 and O1v1. One plasmid was found in all samples known as pC17KP0052-1 and another plasmid with accession no. NZ_CP032191.1 was found only in K90. IncFIB(K) and IncFII(K) are two replicons found in all samples, while ColRNAI replicon was found only in K90. Entero P88, Salmon SEN5, and Klebsi phiKO2 intact phage regions were identified. All samples harbored CRISPR arrays including CRISPR1 and CRISPR2. Our results shed light on critical tasks of mobile genetic elements in ST 627 in antibiotic resistance spreading.

Deciphering Multidrug-Resistant Acinetobacter baumannii from a Pediatric Cancer Hospital in Egypt.

Infection by multidrug-resistant (MDR) Acinetobacter baumannii is one of the major causes of hospital-acquired infections worldwide. The ability of A. baumannii to survive in adverse conditions as well as its extensive antimicrobial resistance make it one of the most difficult to treat pathogens associated with high mortality rates. The aim of this study was to investigate MDR A. baumannii that has spread among pediatric cancer patients in the Children's Cancer Hospital Egypt 57357. Whole-genome sequencing was used to characterize 31 MDR A. baumannii clinical isolates. Phenotypically, the isolates were MDR, with four isolates showing resistance to the last-resort antibiotic colistin. Multilocus sequence typing showed the presence of eight clonal groups, two of which were previously reported to cause outbreaks in Egypt, and one novel sequence type (ST), Oxf-ST2246. Identification of the circulating plasmids showed the presence of two plasmid lineages in the isolates, strongly governed by sequence type. A large number of antimicrobial genes with a range of resistance mechanisms were detected in the isolates, including ß-lactamases and antibiotic efflux pumps. Analysis of insertion sequences (ISs) revealed the presence of ISAba1 and ISAba125 in all the samples, which amplify ß-lactamase expression, causing extensive carbapenem resistance. Mutation analysis was used to decipher underlying mutations responsible for colistin resistance and revealed novel mutations in several outer membrane proteins, in addition to previously reported mutations in pmrB. Altogether, understanding the transmissibility of A. baumannii as well as its resistance and virulence mechanisms will help develop novel treatment options for better management of hospital-acquired infections. IMPORTANCE Acinetobacter baumannii represents a major health threat, in particular among immunocompromised cancer patients. The rise in carbapenem-resistant A. baumannii, and the development of resistance to the last-resort antimicrobial agent colistin, complicates the management of A. baumannii outbreaks and increases mortality rates. Here, we investigate 31 multidrug resistant A. baumannii isolates from pediatric cancer patients in Children's Cancer Hospital Egypt (CCHE) 57357 via whole-genome sequencing. Multilocus sequence typing (MLST) showed the presence of eight clonal groups including a novel sequence type. In silico detection of antimicrobial-resistant genes and virulence factors revealed a strong correlation between certain virulence genes and mortality as well as several point mutations in outer membrane proteins contributing to colistin resistance. Detection of CRISPR/Cas sequences in the majority of the samples was strongly correlated with the presence of prophage sequences and associated with failure of bacteriophage therapy. Altogether, understanding the genetic makeup of circulating A. baumannii is essential for better management of outbreaks.

Author Correction: Genotypic characterization of multiple drug resistant Escherichia coli isolates from a pediatric cancer hospital in Egypt.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Genotypic characterization of multiple drug resistant Escherichia coli isolates from a pediatric cancer hospital in Egypt.

Infection with multiple drug resistant (MDR) Escherichia coli poses a life threat to immunocompromised pediatric cancer patients. Our aim is to genotypically characterize the plasmids harbored in MDR E. coli isolates recovered from bacteremic patients of Children's Cancer Hospital in Egypt 57357 (CCHE 57357). In this study, 21 carbapenem-resistant E. coli (CRE) isolates were selected that exhibit Quinolones and Aminoglycosides resistance. Plasmid shot-gun sequencing was performed using Illumina next- generation sequencing platform. Isolates demonstrated resistant to all beta-lactams, carbapenems, aminoglycosides and quinolones. Of the 32 antimicrobial resistant genes identified that exceeded the analysis cutoff coverage, the highest represented genes were aph(6)-Id, sul2, aph(3″)-Ib, aph(3')-Ia, sul1, dfrA12, TEM-220, NDM-11. Isolates employed a wide array of resistance mechanisms including antibiotic efflux, antibiotic inactivation, antibiotic target replacements and antibiotic target alteration. Sequenced isolates displayed diverse insertion sequences, including IS26, suggesting dynamic reshuffling of the harbored plasmids. Most isolates carried plasmids originating from other bacterial species suggesting a possible horizontal gene transfer. Only two isolates showed virulence factors with iroA gene cluster which was found in only one of them. Outside the realms of nosocomial infections among patients in hospitals, our results indicate a transfer of resistant genes and plasmids across different organisms.