Background
Methods
Patients
Amplification and 454 sequencing
Primer name | 5′–3′ sequencea
| Annealing (°C) |
---|---|---|
SSU-Fwd |
AAGACTCGGCAGCATCTCCAGTGAAATTCTTAGATTTTCTG | 58 |
SSU-Rev |
GCGATCGTCACTGTTCTCCACGTGTTGAGTCAAATTAAGC | |
Cyt-Fwd |
AAGACTCGGCAGCATCTCCAGAGTGGATGGTGTTTTAGAT | 58 |
Cyt-Rev |
GCGATCGTCACTGTTCTCCAGTGCTACCATGTAAATGTAA | |
Clpc-Fwd |
AAGACTCGGCAGCATCTCCAGGTCAATTAACAGAACAA | 55 |
Clpc-Rev |
GCGATCGTCACTGTTCTCCATAGTTAATCTATTTAATAATTC | |
rPLU6b
| TTAAAATTGTTGCAGTTAAAACG | |
rPLU5b
| CCTGTTGTTGCCTTAAACTTC | |
p-AVL-F | GGAATGACAATGTCGTAAAACAAAGTAT | |
P-AVL-R | ATACTTTGTTTTACGACATTGTCATTCC |
Sequence analysis
Plasmodium species | GenBank Acc. |
---|---|
Plasmodium falciparum 18S ribosomal RNA gene (S-type) | HQ283222.1 |
Plasmodium falciparum 18S ribosomal RNA gene (A-type) | JQ627152.1 |
Plasmodium vivax 18S ribosomal RNA gene (A-Type) | JQ627158.1 |
Plasmodium vivax 18S ribosomal RNA gene (O-Type) | U93235.1 |
Plasmodium vivax 18S ribosomal RNA gene (S-Type) | U93234.1 |
Plasmodium berghei 18S ribosomal RNA gene | M19712.1 |
Plasmodium berghei 18S ribosomal RNA gene | M14599.1 |
Plasmodium malariae 18S ribosomal RNA gene | M54897.1 |
Plasmodium ovale 18S ribosomal RNA gene | AB182493.1 |
Plasmodium ovale 18S ribosomal RNA gene | AB182489.1 |
Plasmodium yoelii 18S ribosomal RNA gene | AF180727 |
Plasmodium knowlesi 18S ribosomal RNA gene | JF714686.1 |
Plasmodium cf. inui 18S ribosomal RNA gene | FJ619091.1 |
Plasmodium simiovale 18S ribosomal RNA gene | AB287287.1 |
Plasmodium ovale wallikeri cytochrome b gene | KJ930413.1 |
Plasmodium ovale curtisi cytochrome b gene | KP050432.1 |
Plasmodium malariae cytochrome b gene | LT594637 |
Plasmodium vivax cytochrome b gene | JN788776 |
Plasmodium falciparum cytochrome b gene | KC175316.1 |
Plasmodium berghei cytochrome b gene | DQ414645.1 |
Plasmodium knowlesi cytochrome b gene | JQ345523.1 |
Plasmodium spp. from Gabon cytochrome b gene | AF069623.1 |
Plasmodium yoelii cytochrome b gene | DQ414658.1 |
Plasmodium falciparum HB3 clpC gene for caseinolytic protease C | DQ642846.1 |
Plasmodium ovale curtisi clpC gene for caseinolytic protease C | KP050446.1 |
Plasmodium ovale wallikeri clpC gene for caseinolytic protease C | KP050439 |
Plasmodium malariae clpC gene for caseinolytic protease C | AB649418.1 |
Plasmodium vivax clpC gene for caseinolytic protease C | AB471871.1 |
Plasmodium gaboni clpC gene for caseinolytic protease C | HQ842630.1 |
Plasmodium knowlesi clpCgene for caseinolytic protease C | AB471880.1 |
Plasmodium simiovale clpCgene for caseinolytic protease C | AB471881.1 |
Plasmodium cf. ovale clpC gene for caseinolytic protease C | HQ842632.1 |
Plasmodium yoelii clpC gene for caseinolytic protease C | DQ417625.1 |
Plasmodium berghei clpC gene for caseinolytic protease C | AB649421.1 |
Results
Sequence statistics from patients
Species discrimination by deep sequencing
Species | Infection type | Patients (n) |
---|---|---|
Pfal
| Mono infection | 24 |
Pmal
| Mono infection | 1 |
Pow
| Mono infection | 1 |
Pfal, Pmal
| Double infection | 10 |
Pfal, Poc
| Double infection | 3 |
Pfal, Pow
| Double infection | 2 |
Pfal, Pmal, Pow
| Triple infection | 1 |
Pfal, Poc, Pow
| Triple infection | 1 |
Pfal, Pmal, Poc, Pow
| Quadruple infection | 3 |
Plasmodium falciparum infections
Genotypes | Origin | 386 | 407 | 504 | 517 | 529 | ||
---|---|---|---|---|---|---|---|---|
P. falciparum (JF681166) | China | G | T | G | A | C | ||
P. falciparum (KJ170098) | Gabon | A | C | . | – | . | ||
P. falciparum (KJ170099) | Gabon | . | . | . | . | G | ||
P. falciparum (KJ170100) | Gabon | A | . | . | . | . | ||
P. falciparum (KJ170101) | Gabon | . | . | A | . | . |
1036 | 1189 | |||||||
---|---|---|---|---|---|---|---|---|
P. malariae (AF488000) | Myanmar | G | – | |||||
P. malariae (KJ170105) | Gabon | . | T | |||||
P. malariae (KJ170106) | Gabon | A | T |
1005 | 1026 | 1109 | 1112 | 1128 | 1131 | 1132 | ||
---|---|---|---|---|---|---|---|---|
P. ovale wallikeri (AB182493) | Indonesia | A | C | G | G | A | G | A |
P. ovale wallikeri (KJ170102) | Gabon | . | . | . | . | G | . | . |
P. ovale wallikeri (KJ170103) | Gabon | G | T | A | A | A | G | |
P. ovale wallikeri (KJ170104) | Gabon | G | T | . | . | . | . | . |
Plasmodium malariae infections
Plasmodium ovale wallikeri and P. ovale curtisi infections
Detection of P. ovale curtisi 18S rRNA gene variant
Multiplicity of infection
Sample id | Infecting species | Number of genotypes for each species | ||||
---|---|---|---|---|---|---|
Pfal
|
Pmal
|
Poc
|
Pow
| Total (MOI) | ||
MID01 |
Pmal
| 4 | 4 | |||
MID11 |
Pfal, Pmal
| 2 | 2 | 4 | ||
MID15 |
Pfal
| 2 | 2 | |||
MID16 |
Pfal, Pmal, Poc, Pow
| 1 | 4 | na | 2 | 7 |
MID23 |
Pfal, Poc, Pow
| 2 | 2 | 4 | ||
MID24 |
Pfal, Poc
| 1 | 3 | 4 | ||
MID25 |
Pfal, Pmal
| 1 | 3 | 4 | ||
MID27 |
Pfal, Pmal
| 2 | 2 | 4 | ||
MID28 |
Pfal, Pmal, Poc, Pow
| 2 | 2 | 1 | 1 | 6 |
MID30 |
Pfal, Pmal, Pow
| 1 | na | 2 | 3 | |
MID31 |
Pfal, Poc
| 1 | 3 | 4 | ||
MID32 |
Pfal, Pow
| 1 | 3 | 4 | ||
MID34 |
Pfal, Pmal
| 1 | 2 | 3 | ||
MID38 |
Pfal, Pow
| 1 | 3 | 4 | ||
MID41 |
Pfal, Pmal
| 1 | 2 | 3 | ||
MID43 |
Pfal, Pmal
| 1 | 2 | 3 | ||
MID46 |
Pow
| 2 | 2 |