Comparative Pan-Genomic Analysis Revealed an Improved Multi-Locus Sequence Typing Scheme for Staphylococcus aureus
Abstract
:1. Introduction
2. Materials and Methods
2.1. Sequence Retrieval and S. aureus Genome Data and Gene Annotation
2.2. Pan-Genome Profile Analysis
2.3. Development of an Improved MLST Scheme
2.3.1. Population Structure
2.3.2. New Gene Candidates for S. aureus
2.3.3. Core genes
2.3.4. Role in Cellular Pathways
2.3.5. Mobile Elements
2.3.6. Size of the Gene
2.3.7. Unique Alleles of Each Gene
2.3.8. Data Analysis
Significance of Statistical Parameters
- Tajima’s D value: Tajima’s D value was calculated for already reported MLST scheme and newly predicted genes using “dnasp5”. If the value is zero or closer to zero, it indicates that the observed variation of the population is equal to or closer to the expected. A negative Tajima’s D value shows population expansion. Similarly, a positive value indicates population shrinking or balancing selection. A value between 2 and −2 shows that the genes were driven by neutral selection [31];
- Tajima’s dN/dS ratio: It is a ratio of non-synonymous and synonymous mutations. The ratio is used to distinguish between variants of a population [32];
- dN/dS = (Non-synonymous mutations)/(Synonymous mutations)
2.4. Primers Construction
2.5. Sequence Type Assigning
2.6. MLST Data Analysis
- Based on ST profiles of the genes;
- Based on FASTA sequences of each allele.
3. Results
3.1. Comparative Pan-Genomic Analysis
3.1.1. Staphylococcus aureus Genomic Features and Statistics
3.1.2. Pan-Genome Profile Analysis
3.2. Development of an Improved MLST Scheme
3.2.1. Population Analysis Using Present MLST Scheme
Limitations of Existing Scheme
3.2.2. Selection of New Genes
3.2.3. Statistical Values for New Gene Candidates
New MLST Scheme
Sequence Type Assigning
3.2.4. MLST Tree Construction
4. Discussion
5. Conclusions
Supplementary Materials
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Acknowledgments
Conflicts of Interest
References
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Sr. No. | Gene | Product Size | Left Primer | Tm | GC | Right Primer | Tm | GC |
---|---|---|---|---|---|---|---|---|
1 | opuCC | 682 | CTGGATGTAGTTT GCCCGGA | 59.27 | 55 | TCGCAGC ATAAGGT GGGAAA | 59.63 | 55 |
2 | aspS | 407 | TGTCGTT GCACAAA GTTTAGGT | 58.98 | 45.45 | ATCCTAAT GCAATACC GCCATG | 58.34 | 45 |
3 | rpiB | 419 | GATCCAA ACCGCTC AGCAAA | 59.12 | 50 | GAACCAC GAACTGT TGGACG | 59.42 | 55 |
Sr. No. | Strains | arcC | aroE | glpf | gmk | pta | tpi | yqil | Unreported STs |
---|---|---|---|---|---|---|---|---|---|
1 | FDAARGOS_159 | 4 | 1 | 4 | 12 | 1 | 10 | ||
2 | FDAARGOS_412 | 3 | 32 | 567 | 1 | 4 | 4 | 3 | |
3 | M92 | 2 | 3 | 457 | 1 | 4 | 4 | 3 | |
4 | SA40TW | 19 | 23 | 15 | 2 | 19 | 15 | ||
5 | RF122 | 6 | 72 | 12 | 43 | 49 | 67 | ||
6 | TCH60 | 2 | 2 | 2 | 6 | 3 | 2 | ||
7 | 1971.C01 | 90 | 3 | 1 | 1 | 4 | 4 | 3 | |
8 | 2148.N | 484 | 4 | 1 | 4 | 4 | 3 | ||
9 | NZ15MR0322 | 3 | 19 | 2 | 20 | 26 | 39 | ||
10 | V605 | 2 | 3 | 1 | 238 | 4 | 4 | 3 |
Sr No. | Gene | Process | Size | Unique Alleles | Phi Test | Tajima’s D | dN/dS Ratio |
---|---|---|---|---|---|---|---|
1 | arcC | Amino acid transport and metabolism | 456 | 19/502 | 0.2875 | −2.65316 | 1.19727 |
2 | aroE | Coenzyme transport and metabolism | 456 | 20/502 | 0.2664 | −1.17807 | 12.60328 |
3 | pta | Energy production and conversion | 474 | 19/502 | 0.2723 | −1.80001 | 0.1563 |
4 | Tpi | Energy production and conversion | 402 | 24/502 | 0.2614 | −0.40013 | −1.47932 |
5 | glpF | Carbohydrate transport and metabolism | 465 | 18/502 | 0.2046 | 0.37575 | −0.69466 |
6 | yqil | Inorganic ion transport and metabolism | 516 | 22/502 | 0.9409 | −1.07416 | 0.97535 |
7 | gmk | Carbohydrate transport and metabolism | 417 | 15/502 | 0.5707 | 0.57715 | 4.06854 |
Sr. No | Gene Name | Location | Process | Size (bp) | Unique Alleles |
---|---|---|---|---|---|
1 | ogt | Cytoplasmic | Replication, Recombination, and Repair | 522 | 26/502 |
2 | opuCC | Cell membrane; Lipid-anchor (Probable) [H | Cell wall/membrane biogenesis | 942 | 28/502 |
3 | ureF | Cytoplasmic | Post translation Modification, Protein turnover, Chaperones | 690 | 25/502 |
4 | nrdR | Cytoplasmic | Transcription | 471 | 18/502 |
5 | ureE | Cytoplasmic | Post translation Modification, Protein turnover, Chaperones | 453 | 15/502 |
6 | rpiM | NA | 50S ribosomal protein L13 | 438 | 10/502 |
7 | rpsE | NA | 30S ribosomal protein S5 | 501 | 9/502 |
8 | rplB | NA | 50S ribosomal protein L2 | 834 | 17/502 |
9 | aspS | Cytoplasm | Translation, ribosomal structure, and biogenesis | 1767 | 40/502 |
Sr. No | Name | Phi Test | Invariant Sites | Nucleotide Diversity, Pi | Tajima’s D | Significance | dN/dS Ratio |
---|---|---|---|---|---|---|---|
1 | ogt | × | 0.077346 | 0.02283 | 0.24068 | Not significant, p > 0.10 | 1.03085 |
2 | opuCC | ✔ | 0.043663 | 0.01236 | −2.03506 | *, p < 0.05 | 0.77636 |
3 | ureF | ✔ | 0.021876 | 0.00683 | −2.37067 | p < 0.01 | 0.89544 |
4 | nrdR | ✔ | 0.006988 | 0.00341 | −1.51133 | Not significant, p > 0.10 | 1.34254 |
5 | ureE | ✔ | 0.014496 | 0.00632 | −2.33194 | **, p < 0.01 | 0.91362 |
6 | rpiM | ✔ | 0.003687 | 0.00191 | −1.34752 | Not significant, p > 0.10 | 0.66321 |
7 | rpsE | ✔ | 8.86E−04 | 0.00037 | −1.93483 | p < 0.05 | N/A |
8 | rpiB | ✔ | 0.002087 | 0.00106 | −2.05422 | p < 0.05 | 0.65648 |
9 | aspS | ✔ | 0.021738 | 0.00563 | 1.62031 | 0.10 > p > 0.05 | 0.95247 |
Sr No. | Gene Name | Process | Size | Unique Alleles | Phi Test | Tajima’s D | Non-Syn/Syn Ratio |
---|---|---|---|---|---|---|---|
1 | arc | Amino acid transport and metabolism | 456 | 19/502 | ✔ | −2.6532 | 1.19727 |
2 | Pta | Energy production and conversion | 986 | 19/502 | ✔ | −1.8 | 0.1563 |
3 | Tpi | Energy production and conversion | 368 | 24/502 | ✔ | −0.4001 | −1.4793 |
4 | glpF | Carbohydrate transport and metabolism | 848 | 18/502 | ✔ | 0.37575 | −0.6947 |
5 | opuCC | Cell wall/membrane biogenesis | 942 | 28/502 | ✔ | 0.04366 | −2.0351 |
6 | aspS | Translation, ribosomal structure and biogenesis | 1767 | 40/502 | ✔ | −1.6203 | 0.95247 |
7 | rpiB | 50S ribosomal protein L2 | 834 | 17/502 | ✔ | 0.00209 | −2.0542 |
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Jalil, M.; Quddos, F.; Anwer, F.; Nasir, S.; Rahman, A.; Alharbi, M.; Alshammari, A.; Alshammari, H.K.; Ali, A. Comparative Pan-Genomic Analysis Revealed an Improved Multi-Locus Sequence Typing Scheme for Staphylococcus aureus. Genes 2022, 13, 2160. https://doi.org/10.3390/genes13112160
Jalil M, Quddos F, Anwer F, Nasir S, Rahman A, Alharbi M, Alshammari A, Alshammari HK, Ali A. Comparative Pan-Genomic Analysis Revealed an Improved Multi-Locus Sequence Typing Scheme for Staphylococcus aureus. Genes. 2022; 13(11):2160. https://doi.org/10.3390/genes13112160
Chicago/Turabian StyleJalil, Maira, Fatima Quddos, Farha Anwer, Samavi Nasir, Abdur Rahman, Metab Alharbi, Abdulrahman Alshammari, Huda Kamel Alshammari, and Amjad Ali. 2022. "Comparative Pan-Genomic Analysis Revealed an Improved Multi-Locus Sequence Typing Scheme for Staphylococcus aureus" Genes 13, no. 11: 2160. https://doi.org/10.3390/genes13112160
APA StyleJalil, M., Quddos, F., Anwer, F., Nasir, S., Rahman, A., Alharbi, M., Alshammari, A., Alshammari, H. K., & Ali, A. (2022). Comparative Pan-Genomic Analysis Revealed an Improved Multi-Locus Sequence Typing Scheme for Staphylococcus aureus. Genes, 13(11), 2160. https://doi.org/10.3390/genes13112160