Synonymous Mutations in rpsT Lead to Ribosomal Assembly Defects That Can Be Compensated by Mutations in fis and rpoA

Abstract Background Red blood cell invasion by the Plasmodium vivax merozoite requires interaction between the Duffy antigen receptor for chemokines (DARC) and the P. vivax Duffy-binding protein II (PvDBPII). Given that the disruption of this interaction prevents P. vivax blood-stage infection, a PvDBP-based vaccine development has been well recognized. However, the polymorphic nature of PvDBPII prevents a strain transcending immune response and complicates attempts to design a vaccine. Methods Twenty-three P. vivax clinical isolates collected from three areas of Ethiopia were sequenced at the pvdbpII locus. A total of 392 global pvdbpII sequences from seven P. vivax endemic countries were also retrieved from the NCBI archive for comparative analysis of genetic diversity, departure from neutrality, linkage disequilibrium, genetic differentiation, PvDBP polymorphisms, recombination and population structure of the parasite population. To establish a haplotype relationship a network was constructed using the median joining algorithm. Results A total of 110 variable sites were found, of which 44 were parsimony informative. For Ethiopian isolates there were 12 variable sites of which 10 were parsimony informative. These parsimony informative variants resulted in 10 nonsynonymous mutations. The overall haplotype diversity for global isolates was 0.9596; however, the haplotype diversity was 0.874 for Ethiopia. Fst values for genetic revealed Ethiopian isolates were closest to Indian isolates as well as to Sri Lankan and Sudanese isolates but further away from Mexican, Papua New Guinean and South Korean isolates. There was a total of 136 haplotypes from the 415 global isolates included for this study. Haplotype prevalence ranged from 36.76% to 0.7%, from this 74.2% were represented by single parasite isolates. None of the Ethiopian isolates grouped with the Sal I reference haplotype. From the total observed nonsynonymous mutations 13 mapped to experimentally verified epitope sequences. Including 10 non-synonymous mutations from Ethiopia. However, all the polymorphic regions in Ethiopian isolates were located away from DARC, responsible for junction formation. Conclusion The results of this study are concurrent with the multivalent vaccine approach to design an effective treatment. However, the presence of novel haplotypes in Ethiopian isolates that were not shared by other global sequences warrant further investigation.

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Pseudomonas aeruginosa PA80 is a cystic fibrosis isolate deficient in RhlRI quorum sensing

Scientific Reports ◽

10.1038/s41598-021-85100-0 ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Syed A. K. Shifat Ahmed ◽

Michelle Rudden ◽

Sabrina M. Elias ◽

Thomas J. Smyth ◽

Roger Marchant ◽

...

Keyword(s):

Cystic Fibrosis ◽

Pseudomonas Aeruginosa ◽

Quorum Sensing ◽

Virulence Factors ◽

Genomic Islands ◽

Clinical Strain ◽

Whole Genome ◽

Synonymous Mutations ◽

Wide Range

AbstractPseudomonas aeruginosa uses quorum sensing (QS) to modulate the expression of several virulence factors that enable it to establish severe infections. The QS system in P. aeruginosa is complex, intricate and is dominated by two main N-acyl-homoserine lactone circuits, LasRI and RhlRI. These two QS systems work in a hierarchical fashion with LasRI at the top, directly regulating RhlRI. Together these QS circuits regulate several virulence associated genes, metabolites, and enzymes in P. aeruginosa. Paradoxically, LasR mutants are frequently isolated from chronic P. aeruginosa infections, typically among cystic fibrosis (CF) patients. This suggests P. aeruginosa can undergo significant evolutionary pathoadaptation to persist in long term chronic infections. In contrast, mutations in the RhlRI system are less common. Here, we have isolated a clinical strain of P. aeruginosa from a CF patient that has deleted the transcriptional regulator RhlR entirely. Whole genome sequencing shows the rhlR locus is deleted in PA80 alongside a few non-synonymous mutations in virulence factors including protease lasA and rhamnolipid rhlA, rhlB, rhlC. Importantly we did not observe any mutations in the LasRI QS system. PA80 does not appear to have an accumulation of mutations typically associated with several hallmark pathoadaptive genes (i.e., mexT, mucA, algR, rpoN, exsS, ampR). Whole genome comparisons show that P. aeruginosa strain PA80 is closely related to the hypervirulent Liverpool epidemic strain (LES) LESB58. PA80 also contains several genomic islands (GI’s) encoding virulence and/or resistance determinants homologous to LESB58. To further understand the effect of these mutations in PA80 QS regulatory and virulence associated genes, we compared transcriptional expression of genes and phenotypic effects with isogenic mutants in the genetic reference strain PAO1. In PAO1, we show that deletion of rhlR has a much more significant impact on the expression of a wide range of virulence associated factors rather than deletion of lasR. In PA80, no QS regulatory genes were expressed, which we attribute to the inactivation of the RhlRI QS system by deletion of rhlR and mutation of rhlI. This study demonstrates that inactivation of the LasRI system does not impact RhlRI regulated virulence factors. PA80 has bypassed the common pathoadaptive mutations observed in LasR by targeting the RhlRI system. This suggests that RhlRI is a significant target for the long-term persistence of P. aeruginosa in chronic CF patients. This raises important questions in targeting QS systems for therapeutic interventions.

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Functional alterations caused by mutations reflect evolutionary trends of SARS-CoV-2

Briefings in Bioinformatics ◽

10.1093/bib/bbab042 ◽

2021 ◽

Author(s):

Liang Cheng ◽

Xudong Han ◽

Zijun Zhu ◽

Changlu Qi ◽

Ping Wang ◽

...

Keyword(s):

Reference Genome ◽

Sequence Data ◽

Purifying Selection ◽

Virus Genome ◽

Receptor Binding Domain ◽

Evolutionary Trends ◽

Synonymous Mutations ◽

Almost All ◽

Virus Strains ◽

New Mutations

Abstract Since the first report of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in December 2019, the COVID-19 pandemic has spread rapidly worldwide. Due to the limited virus strains, few key mutations that would be very important with the evolutionary trends of virus genome were observed in early studies. Here, we downloaded 1809 sequence data of SARS-CoV-2 strains from GISAID before April 2020 to identify mutations and functional alterations caused by these mutations. Totally, we identified 1017 nonsynonymous and 512 synonymous mutations with alignment to reference genome NC_045512, none of which were observed in the receptor-binding domain (RBD) of the spike protein. On average, each of the strains could have about 1.75 new mutations each month. The current mutations may have few impacts on antibodies. Although it shows the purifying selection in whole-genome, ORF3a, ORF8 and ORF10 were under positive selection. Only 36 mutations occurred in 1% and more virus strains were further analyzed to reveal linkage disequilibrium (LD) variants and dominant mutations. As a result, we observed five dominant mutations involving three nonsynonymous mutations C28144T, C14408T and A23403G and two synonymous mutations T8782C, and C3037T. These five mutations occurred in almost all strains in April 2020. Besides, we also observed two potential dominant nonsynonymous mutations C1059T and G25563T, which occurred in most of the strains in April 2020. Further functional analysis shows that these mutations decreased protein stability largely, which could lead to a significant reduction of virus virulence. In addition, the A23403G mutation increases the spike-ACE2 interaction and finally leads to the enhancement of its infectivity. All of these proved that the evolution of SARS-CoV-2 is toward the enhancement of infectivity and reduction of virulence.

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1456. Resistance Mechanisms of Tigecycline in Enterococcus faecalis

Open Forum Infectious Diseases ◽

10.1093/ofid/ofaa439.1637 ◽

2020 ◽

Vol 7 (Supplement_1) ◽

pp. S730-S731

Author(s):

Bing Bai ◽

Zewen Wen ◽

Zhiwei Lin ◽

Tam Vincent H ◽

Zhijian Yu

Keyword(s):

Enterococcus Faecalis ◽

Molecular Mechanisms ◽

Genetic Mutations ◽

Dilution Method ◽

Agar Dilution Method ◽

Whole Genome ◽

Synonymous Mutations ◽

Target Sites ◽

Parental Isolate ◽

Resistant Strains

Abstract Background Enterococcus faecalis have been regarded as one of the leading causes of the nosocomial infections worldwide. Tigecycline (TGC) is considered as a choice of last resort for the treatment of infections caused by multidrug-resistant E. faecalis, however, the emergence of TGC non-susceptibility has posted the therapeutic challenge. Non-susceptibility in clinical strains could be due to resistance (MIC >0.5 mg/l) or heteroresistance. Therefore, this study aimed to understand the underlying molecular mechanisms of TGC resistance and heteroresistance in E. faecalis. Methods In vitro induction experiments were carried out under TGC pressure with two TGC- sensitive E. faecalis strains. Heteroresistance was evaluated by population analysis profiling (PAP) in 270 clinical TGC- sensitive E. faecalis strains. TGC susceptibility was determined by the agar dilution method. Resistance and heteroresistance mechanisms were investigated by identifying genetic mutations in tetracycline (Tet) target sites and susceptibility testing in the presence of the efflux protein inhibitors phenylalanine-arginine-β-naphthylamide (PaβN) and carbonyl cyanide m chlorophenylhydrazine (CCCP). Comparison of single nucleotide polymorphism in the whole genome between the parental isolate and two TGC-resistant strains were investigated by next-generation sequencing. Results No mutations in Tet target sites in seven TGC heteroresistant strains were present, whereas the mutations in Tet target sites of seven TGC-resistant E. faecalis were frequently found (Table 1). TGC MICs in heteroresistant strains were reduced by CCCP (Table 2). Whole genome sequencing revealed the same non-synonymous mutations and transcoding deletions in the exons of several genes encoding for various enzymes or transfer systems (Table 3). Table 1. The characteristics of the antimicrobial susceptibility, resistance mechanism of TGC-induced resistant isolates Table 2. Characteristics of clinical heteroresistant mother E. faecalis strains and heteroresistance-derived E. faecalis clones Table 3. List of mutation-related genes, amino acids and proteins by comparison of whole genome between the parental isolate and the TGC-induced resistant strains Conclusion Our data indicated that the main mechanism of TGC heteroresistance in E. faecalis might be associated with the efflux pumps. TGC resistance in E. faecalis was associated with mutations in the 16SrRNA site or 30S ribosome protein S10. The genetic mutations in several enzymes and transfer systems might also participate in the resistance development to TGC in E. faecalis. Disclosures All Authors: No reported disclosures

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Virus Adaptation Following Experimental Infection of Chickens with a Domestic Duck Low Pathogenic Avian Influenza Isolate from the 2017 USA H7N9 Outbreak Identifies Polymorphic Mutations in Multiple Gene Segments

Viruses ◽

10.3390/v13061166 ◽

2021 ◽

Vol 13 (6) ◽

pp. 1166

Author(s):

Klaudia Chrzastek ◽

Karen Segovia ◽

Mia Torchetti ◽

Mary Lee Killian ◽

Mary Pantin-Jackwood ◽

...

Keyword(s):

Avian Influenza ◽

Experimental Infection ◽

Clinical Signs ◽

Interspecies Transmission ◽

Clinical Samples ◽

Domestic Duck ◽

Receptor Binding Site ◽

Synonymous Mutations ◽

Field Samples ◽

Virus Adaptation

In March 2017, highly pathogenic (HP) and low pathogenic (LP) avian influenza virus (AIV) subtype H7N9 were detected from poultry farms and backyard birds in several states in the southeast United States. Because interspecies transmission is a known mechanism for evolution of AIVs, we sought to characterize infection and transmission of a domestic duck-origin H7N9 LPAIV in chickens and genetically compare the viruses replicating in the chickens to the original H7N9 clinical field samples used as inoculum. The results of the experimental infection demonstrated virus replication and transmission in chickens, with overt clinical signs of disease and shedding through both oral and cloacal routes. Unexpectedly, higher levels of virus shedding were observed in some cloacal swabs. Next generation sequencing (NGS) analysis identified numerous non-synonymous mutations at the consensus level in the polymerase genes (i.e., PA, PB1, and PB2) and the hemagglutinin (HA) receptor binding site in viruses recovered from chickens, indicating possible virus adaptation in the new host. For comparison, NGS analysis of clinical samples obtained from duck specimen collected during the outbreak indicated three polymorphic sides in the M1 segment and a minor population of viruses carrying the D139N (21.4%) substitution in the NS1 segment. Interestingly, at consensus level, A/duck/Alabama (H7N9) had isoleucine at position 105 in NP protein, similar to HPAIV (H7N9) but not to LPAIV (H7N9) isolated from the same 2017 influenza outbreak in the US. Taken together, this work demonstrates that the H7N9 viruses could readily jump between avian species, which may have contributed to the evolution of the virus and its spread in the region.

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The Age of Nonsynonymous and Synonymous Mutations in Animal mtDNA and Implications for the Mildly Deleterious Theory

Genetics ◽

10.1093/genetics/153.1.497 ◽

1999 ◽

Vol 153 (1) ◽

pp. 497-506 ◽

Cited By ~ 4

Author(s):

Rasmus Nielsen ◽

Daniel M Weinreich

Keyword(s):

Dna Sequences ◽

Purifying Selection ◽

Data Sets ◽

Deleterious Mutations ◽

Synonymous Mutations ◽

Weak Evidence ◽

Mitochondrial Data ◽

The Mean ◽

Excess Number ◽

Neutral Mutations

Abstract McDonald/Kreitman tests performed on animal mtDNA consistently reveal significant deviations from strict neutrality in the direction of an excess number of polymorphic nonsynonymous sites, which is consistent with purifying selection acting on nonsynonymous sites. We show that under models of recurrent neutral and deleterious mutations, the mean age of segregating neutral mutations is greater than the mean age of segregating selected mutations, even in the absence of recombination. We develop a test of the hypothesis that the mean age of segregating synonymous mutations equals the mean age of segregating nonsynonymous mutations in a sample of DNA sequences. The power of this age-of-mutation test and the power of the McDonald/Kreitman test are explored by computer simulations. We apply the new test to 25 previously published mitochondrial data sets and find weak evidence for selection against nonsynonymous mutations.

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Kelch 13-propeller polymorphisms in Plasmodium falciparum from Jazan region, southwest Saudi Arabia

Malaria Journal ◽

10.1186/s12936-020-03467-3 ◽

2020 ◽

Vol 19 (1) ◽

Author(s):

Ommer Mohammed Dafalla ◽

Mohammed Alzahrani ◽

Ahmed Sahli ◽

Mohammed Abdulla Al Helal ◽

Mohammad Mohammad Alhazmi ◽

...

Keyword(s):

Plasmodium Falciparum ◽

Saudi Arabia ◽

Parasite Clearance ◽

Sub Saharan Africa ◽

Local Alignment ◽

Nucleotide Polymorphisms ◽

Gene Encoding ◽

Synonymous Mutations ◽

Search Tool ◽

Sub Saharan

Abstract Background Artemisinin-based combination therapy (ACT) is recommended at the initial phase for treatment of Plasmodium falciparum, to reduce morbidity and mortality in all countries where malaria is endemic. Polymorphism in portions of P. falciparum gene encoding kelch (K13)-propeller domains is associated with delayed parasite clearance after ACT. Of about 124 different non-synonymous mutations, 46 have been identified in Southeast Asia (SEA), 62 in sub-Saharan Africa (SSA) and 16 in both the regions. This is the first study designed to analyse the prevalence of polymorphism in the P. falciparum k13-propeller domain in the Jazan region of southwest Saudi Arabia, where malaria is endemic. Methods One-hundred and forty P. falciparum samples were collected from Jazan region of southwest Saudi Arabia at three different times: 20 samples in 2011, 40 samples in 2016 and 80 samples in 2020 after the implementation of ACT. Plasmodium falciparum kelch13 (k13) gene DNA was extracted, amplified, sequenced, and analysed using a basic local alignment search tool (BLAST). Results This study obtained 51 non-synonymous (NS) mutations in three time groups, divided as follows: 6 single nucleotide polymorphisms (SNPs) ‘11.8%’ in samples collected in 2011 only, 3 (5.9%) in 2011and 2016, 5 (9.8%) in 2011 and 2020, 5 (9.8%) in 2016 only, 8 (15.7%) in 2016 and 2020, 14 (27.5%) in 2020 and 10 (19.6%) in all the groups. The BLAST revealed that the 2011 isolates were genetically closer to African isolates (53.3%) than Asian ones (46.7%). Interestingly, this proportion changed completely in 2020, to become closer to Asian isolates (81.6%) than to African ones (18.4%). Conclusions Despite the diversity of the identified mutations in the k13-propeller gene, these data did not report widespread artemisinin-resistant polymorphisms in the Jazan region where these samples were collected. Such a process would be expected to increase frequencies of mutations associated with the resistance of ACT.

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Correlation between ZBRK1/ZNF350 gene polymorphism and breast cancer

BMC Medical Genomics ◽

10.1186/s12920-020-00862-2 ◽

2021 ◽

Vol 14 (1) ◽

Author(s):

Jun Wu ◽

Alibiati Eni ◽

Eliar Roussuri ◽

Binlin Ma

Keyword(s):

Breast Cancer ◽

Family History ◽

Gene Polymorphism ◽

Zinc Finger ◽

Zinc Finger Protein ◽

Control Groups ◽

Wild Genotype ◽

Mutant Genotype ◽

Synonymous Mutations ◽

Significant Difference

Abstract Background This study is to explore the relationship between the ZBRK1/ZNF350 (Zinc finger and BRCA1-interacting protein with KRAB domain-1; also known as zinc-finger protein 350) gene polymorphism and early-onset breast cancer. Methods The ZBRK1/ZNF350 gene exon detection analysis was performed with the direct sequencing and Snapshot methods in 80 cases of breast cancer (aged ≤ 40 years old) and 240 healthy subjects (aged ≤ 40 years old). Results Totally 9 sequence variants were detected, including 5 missense mutations and 4 synonymous mutations, located at EXON3, EXON4 and EXON5, respectively. The rs4987241 and rs3764538 variants were published for the first time, while the remaining variants had been reported before. There were significant differences in the frequency distribution of family history between the breast cancer and control groups. Moreover, there were significant differences in the CT genotype frequency at the rs138898320 locus between the breast cancer and healthy control groups. Compared with the carriers of CC wild genotype at rs138898320, the risk of breast cancer was reduced by 88.3% in the CT mutant genotype carriers, with significant difference. In the stratification with no family history, compared with the carriers of CC wild genotype at rs138898320, significant differences were observed for the CT mutant genotype carriers. In the stratification with family history, there was no significant difference in the variation of rs138898320. Conclusion The rs138898320 CT mutation genotype of ZBRK1/ZNF350 may reduce the risk of breast cancer, and the protecting effect would be increased in the stratification with no family history. Trial registration Not applicable.

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Genetic stability of genome-scale deoptimized RNA virus vaccine candidates under selective pressure

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1619242114 ◽

2017 ◽

Vol 114 (3) ◽

pp. E386-E395 ◽

Cited By ~ 26

Author(s):

Cyril Le Nouën ◽

Thomas McCarty ◽

Michael Brown ◽

Melissa Laird Smith ◽

Roberto Lleras ◽

...

Keyword(s):

Virus Vaccine ◽

Genetic Stability ◽

Selective Pressure ◽

Rna Virus ◽

Temperature Sensitive ◽

Phenotypic Analysis ◽

Vaccine Candidates ◽

Synonymous Mutations ◽

Genome Scale

Recoding viral genomes by numerous synonymous but suboptimal substitutions provides live attenuated vaccine candidates. These vaccine candidates should have a low risk of deattenuation because of the many changes involved. However, their genetic stability under selective pressure is largely unknown. We evaluated phenotypic reversion of deoptimized human respiratory syncytial virus (RSV) vaccine candidates in the context of strong selective pressure. Codon pair deoptimized (CPD) versions of RSV were attenuated and temperature-sensitive. During serial passage at progressively increasing temperature, a CPD RSV containing 2,692 synonymous mutations in 9 of 11 ORFs did not lose temperature sensitivity, remained genetically stable, and was restricted at temperatures of 34 °C/35 °C and above. However, a CPD RSV containing 1,378 synonymous mutations solely in the polymerase L ORF quickly lost substantial attenuation. Comprehensive sequence analysis of virus populations identified many different potentially deattenuating mutations in the L ORF as well as, surprisingly, many appearing in other ORFs. Phenotypic analysis revealed that either of two competing mutations in the virus transcription antitermination factor M2-1, outside of the CPD area, substantially reversed defective transcription of the CPD L gene and substantially restored virus fitness in vitro and in case of one of these two mutations, also in vivo. Paradoxically, the introduction into Min L of one mutation each in the M2-1, N, P, and L proteins resulted in a virus with increased attenuation in vivo but increased immunogenicity. Thus, in addition to providing insights on the adaptability of genome-scale deoptimized RNA viruses, stability studies can yield improved synthetic RNA virus vaccine candidates.

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Highly conserved gsc1 gene of Pneumocystis jirovecii in patients with or without prior exposure to Echinocandins

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.01563-21 ◽

2021 ◽

Author(s):

Pierre L. Bonnet ◽

Solène Le Gal ◽

Claire V. Hoffmann ◽

Florent Morio ◽

Fouleymata Diabira ◽

...

Keyword(s):

Hot Spots ◽

Selection Pressure ◽

Gene Diversity ◽

Prior Exposure ◽

Cell Wall Component ◽

Synonymous Mutations ◽

Significant Difference ◽

A Cell ◽

Patient Groups ◽

Noncompetitive Inhibitors

Echinocandins are noncompetitive inhibitors of the GSC1 subunit of the enzymatic complex involved in synthesis of 1,3-beta-D-glucan, a cell wall component of most fungi, including Pneumocystis spp. Echinocandins are widely used for treating systemic candidiasis and rarely used for treating Pneumocystis pneumonia. Consequently, data on P. jirovecii gsc1 gene diversity are still scarce, compared to the homologous fks1 gene of Candida spp. In this study, we analyzed P. jirovecii gsc1 gene diversity and the putative selection pressure of echinocandins on P. jirovecii. Gsc1 gene sequences of P. jirovecii specimens from two patient groups were compared. One group of 27 patients had prior exposure to echinocandins whereas the second group of 24 patients did not, at the time of P. jirovecii infection diagnoses. Two portions of P. jirovecii gsc1 gene, HS1 and HS2, homologous to hot spots described in Candida spp., were sequenced. Three SNPs at positions 2204, 2243, and 2303 close to the HS1 region and another SNP at position 4540 more distant from the HS2 region were identified. These SNPs represent synonymous mutations. Three gsc1 HS1 alleles, A, B, and C, and two gsc1 HS2 alleles, a and b, and four haplotypes, Ca, Cb, Aa, and Ba, were defined, without significant difference in haplotype distribution in both patient groups ( p = 0.57). Considering the identical diversity of P. jirovecii gsc1 gene and the detection of synonymous mutations in both patient groups, no selection pressure of echinocandins among P. jirovecii microorganisms can be pointed out so far.

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