Sir,

Flaviviruses (FVs) are arthropod-borne viruses that affect humans in tropical regions worldwide.\[[@ref1]\] Previous reports\[[@ref1][@ref2]\] have shown the importance of implementing FV novel detection strategies in tropical regions, such as the use of universal primers that enable a simultaneous identification of these pathogens in a single polymerase chain reaction (PCR).\[[@ref1][@ref2][@ref3]\]

We analyzed 462 blood samples from febrile patients in western Mexico. FV identification was carried out through a PCR assay using universal primers validated *in vitro* by Maher-Sturgess *et al*. A total of 74 blood samples amplified with different amplicon sizes similar to those previously described.\[[@ref4]\] Our DNA sequencing analysis showed the identification of dengue virus serotype 1 in 31 blood samples,\[[@ref5]\] and the remaining 43 amplicons corresponded to different human gene regions \[[Table 1](#T1){ref-type="table"}\], representing 9.3% of all the samples analyzed.

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Polymerase chain reaction results using the universal primers, Flav100F and Flav200R, corresponding to different sizes of amplicons of human genes

  Number of samples   Description                                                                                                                         Query position    Amplicon length (bp)   Query cover (%)   *E*   Accession number
  ------------------- ----------------------------------------------------------------------------------------------------------------------------------- ----------------- ---------------------- ----------------- ----- ------------------
  8                   *Homo sapiens* mitochondrial ND1 gene\*                                                                                             191-867           \~690                  \~98              0.0   LC178870.1
  1                   *Homo sapiens* isolate XEB057 mitochondrion                                                                                         3496-4236         758                    97                0.0   KF056243.1
  1                   *Homo sapiens* chromosome 15 clone RP11-353B9 map 15q21                                                                             67,269-67,987     735                    97                0.0   AC018927.6
  1                   *Homo sapiens* ELP4                                                                                                                 175,278-174,496   796                    98                0.0   NG_034086.1
  1                   *Homo sapiens* isolate GranChaco16 haplogroup B2 mitochondrion                                                                      3496-4172         691                    97                0.0   KP172434.1
  1                   *Homo sapiens* EHBP1                                                                                                                195,453-196,134   704                    96                0.0   NG_016758.1
  1                   *Homo sapiens* ROBO1                                                                                                                282,692-283,334   651                    98                0.0   NG_011729.1
  1                   *Homo sapiens* DPYD                                                                                                                 768,218-768,819   622                    96                0.0   NG_008807.2
  1                   *Homo sapiens* chromosome 8, clone RP11-449M6                                                                                       109,136-109,669   553                    96                0.0   AC090574.6
  1                   *Homo sapiens* FAM189A2                                                                                                             39,721-403,20     615                    97                0.0   NG_052888.1
  1                   *Homo sapiens* isolate HGDP00234 mitochondrion                                                                                      3496-4171         693                    97                0.0   KP240920.1
  1                   *Homo sapiens* SGCD                                                                                                                 626,147-626,990   851                    99                0.0   NG_008693.2
  1                   Human DNA sequence from clone RP11-138I18 on chromosome 10                                                                          20,188-21,041     871                    98                0.0   AL512631.11
  1                   *Homo sapiens* isolate mtGHispPR0007 mitochondrion                                                                                  3492-4168         691                    97                0.0   KM102107.1
  1                   *Homo sapiens* chromosome 1 clone RP11-84O12                                                                                        137,829-138,704   893                    98                0.0   AC099670.2
  1                   Human DNA sequence from clone GS1-115G20 on chromosome 1                                                                            148,903-149,647   764                    97                0.0   AL078645.31
  1                   Human DNA sequence from clone RP11-541G7 on chromosome X                                                                            31,561-32,248     706                    97                0.0   AL590410.13
  1                   *Homo sapiens* apolipoprotein L, 3, mRNA                                                                                            798-1432          649                    97                0.0   BC042918.1
  1                   *Homo sapiens* ERBB4                                                                                                                532,004-532,840   844                    99                0.0   NG_011805.1
  1                   *Homo sapiens* genomic DNA, chromosome 11q, clone: CMB9-14B22                                                                       60,374-60,956     594                    98                0.0   AP000484.5
  1                   *Homo sapiens* steroid-5-alpha-reductase, alpha polypeptide 1 (3-oxo-5 alpha-steroid delta 4-dehydrogenase alpha 1) (SRD5A1) gene   23,462-24,192     751                    97                0.0   AY341029.1
  1                   *Homo sapiens* BAC clone RP11-421J10                                                                                                15,356-15,933     602                    96                0.0   AC009236.4
  1                   *Homo sapiens* isolate HGDP00234 mitochondrion                                                                                      3495-4171         688                    98                0.0   KP240920.1
  1                   *Homo sapiens* FAM189A2, RefSeqGene on chromosome 9                                                                                 39,721-40,318     615                    97                0.0   NG_052888.1
  1                   Human DNA sequence from clone RP11-420G9 on chromosome 1                                                                            1532-1905         385                    97                0.0   AL358013.19
  1                   Human DNA sequence from clone RP13-766B11 on chromosome X                                                                           18,172-18,595     434                    97                0.0   BX510661.4
  1                   *Homo sapiens* BAC clone RP11-717F1 from 16                                                                                         1021-1567         558                    98                0.0   AC079801.2
  1                   Human DNA sequence from clone RP11-327L9 on chromosome 9                                                                            34,895-35,445     567                    97                0.0   AL161782.14
  1                   *Homo sapiens* BAC clone RP11-354N12 from 2                                                                                         80,765-81,132     381                    96                0.0   AC009960.10
  1                   *Homo sapiens* LAMA2                                                                                                                12,902-13,384     502                    96                0.0   NG_008678.1
  1                   Human DNA sequence from clone RP11-267I18 on chromosome 13                                                                          60,062-60,535     490                    96                0.0   AL162852.14
  1                   Human DNA sequence from clone RP11-49A19 on chromosome 13                                                                           20,879-21,452     582                    98                0.0   AL354741.16
  1                   *Homo sapiens* chromosome 1 clone RP11-335E6                                                                                        71,818-72,229     431                    95                0.0   AC096543.2
  1                   *Homo sapiens* isolate mtGHispTX0026 mitochondrion                                                                                  3496-3870         386                    97                0.0   KM102130.1
  1                   *Homo sapiens* 3 BAC RP11-561L3 (Roswell Park Cancer Institute Human BAC Library)                                                   16,361-16,723     376                    96                0.0   AC092969.7
  1                   *Homo sapiens* UTRN, RefSeqGene on chromosome 6                                                                                     22,2195-222,599   424                    95                0.0   NG_042293.1

\*Sequence analysis of these amplified segments showed a similarity variation of \~4 bp. ELP4: Elongator acetyltransferase complex subunit 4, EHBP1: EH domain-binding protein 1, ROBO1: Roundabout guidance receptor 1, DPYD: Dihydropyrimidine dehydrogenase, FAM189A2: Family with sequence similarity 189 member A2, SGCD: Sarcoglycan delta, ERBB4: Erb-B2 receptor tyrosine kinase 4, LAMA2: Laminin subunit alpha 2, UTRN: Utrophin

Undoubtedly, the use of nonvalidated universal PCR primers in natural hosts represents a risk for obtaining unreliable results in animal and human populations due to the diversity of genomes. The development of universal PCR primers for FV detection is a potential tool for its diagnosis, especially in tropical regions where FVs frequently coexist, so febrile symptoms associated with FVs easily could result in a diagnostic confusion. This clinical confusion could interfere on the deployment of effective solutions for the detection, prevention, and control of emergent FV outbreaks in timely manner; therefore, the control of FV diseases could be hampered if effective and reliable detection strategies are not implemented.

Financial support and sponsorship {#sec2-1}
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This work was supported by the National Council for Science and Technology of Mexico (CONACYT-FORDECyT grant \#2009/1-117535).

Conflicts of interest {#sec2-2}
=====================

There are no conflicts of interest.

The authors would like to thank the National Council for Science and Technology (Federal Government, Mexico) for the financial assistance provided through CONACYT-FORDECyT grant \#2009/1-117535.
