# Human gyrovirus in healthy blood donors, france.
Philippe Biagini, Sandra Bédarida, Mhammed Touinssi, Vital Galicher

► To cite this version:

Philippe Biagini, Sandra Bédarida, Mhammed Touinssi, Vital Galicher. Human gyrovirus in healthy blood donors, france. Emerging Infectious Diseases, 2013, 19 (6), pp.1014-5.
10.3201/eid1906.130228 hal-00831443

HAL Id: hal-00831443

https://hal.science/hal-00831443

Submitted on 12 Oct 2023

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# Human Gyrovirus in Healthy Blood Donors, France

**To the Editor:** Gyroviruses (GyVs) are naked, single-stranded DNA viruses that were described in chickens in 1979 (1). Chicken anemia virus (CAV), initially the sole member of the genus *Gyrovirus* (family *Circoviridae*), possesses a genome of ≈2.3 kb, containing 3 major partially overlapping open reading frames, viral proteins [VP] 1–3, and a short untranslated region (1). For >30 years, this virus, which was responsible for severe anemia and increased death rates in young chickens, was considered to have an extremely low genetic diversity and to be specific to this animal host. In 2011, however, sequence-independent molecular protocols enabled the characterization of highly divergent, GyV-related sequences in human and chicken biological samples. Human GyV 1 (HGyV1), avian GyV 2, and GyV3 sequences were identified from human skin, chicken blood, and human feces, respectively (2–4). These genomes harbor a genetic organization similar to CAV, despite a high genetic divergence (49%–65%).

One study described the detection of GyVs in HIV-positive patients and kidney transplant recipients (0.7% and 6%, respectively) (5), but these viruses had not been identified in blood samples from healthy persons. We investigated the presence of HGyV DNA in 352 blood samples from healthy blood

donors in France (mean age 39 years; 185 men; M:F ratio 1:1.11).

Plasma samples were prepared as described (6), and 1-mL aliquots were used for nucleic acids extraction (MagNA pure LC; Roche Diagnostics, Meylan, France). HGyV DNA was detected by using 2 systems in separate real-time TaqMan amplification assays (StepOne Plus; Applied Biosystems, Courto-boeuf, France). The first detection assay (VP1 gene) was described previously (HGyV-rtFP/HGyV-rtRP primers, HGyV-rtP probe, 72 nt) (5). The second assay was designed following the analysis of available HGyV sequences (Figure): sense primer HGyVsPBs 5'-GCTAAGACTGTRACATGGC-3', reverse primer HGyVsPBr 5'-CTC-CGGGAATAGCGTCTTC-3', probe HGyVsPBp 5'-FAM-TGGCACTGGAGACACAGACTGCG-TAM-RA-3'. This assay targets the VP2 gene of the viral genome, with an expected length of 118–115 bp, depending on the reference sequence considered.

Amplification reactions were performed by using 10% of extracted material with the TaqMan Fast Universal PCR Kit (Applied Biosystems) in a final volume of 20 µL. Cycling conditions for both assays were 95°C for 20 s, followed by 50 cycles of 95°C for 1 s and 60°C for 20 s. The sensitivity of TaqMan assays was estimated to be 10 copies of HGyV DNA by using dilutions of a synthetic template. Each amplification product was subjected to additional agarose gel electrophoresis to help eliminate potential false-negative real-time PCR results.

Among the 352 plasma samples tested, 3 (0.85%) resulted in a positive signal by using our in-house real-time detection assay; no positive signal was identified by using the other system tested. When the tests were repeated, identical results were obtained. No additional amplicons were identifiable after agarose gel analysis. HGyV DNA titers in the 3 positive samples were low (<500 copies/mL plasma). Positive blood samples originated from 1 woman (age 44) and 2 men (ages 29 and 39). No biological or serologic marker evaluated for routine blood donor screening in France was associated with these donations.

Partial HGyV sequences obtained were cloned and sequenced (7 clones each). All sequences characterized clustered with HGyV1 sequences, exhibiting either 100% nucleotide identity or 1 point mutation (corresponding to a nonsynonymous substitution, ala → thr) (Figure). No intragenetic diversity was identified.

Our results demonstrate that recently discovered HGyVs are detectable in blood of healthy persons. The low prevalence (0.85%) suggests, however, that the virus is infrequently found in the general population in France. A study from Italy of HGyVs in blood from healthy donors did not detect such viruses (5). This result may be linked to the small number of samples tested (n = 50) or to the use of a nonoptimized detection system designed on the basis of HGyV1 sequences only; moreover, the possibility that the VP1 and VP2 amplified

9B1 acatggcAAGACTACGAAGAAGACGACCTCGCGGACGCTTTGGCTTCTACCACAGAGGACGATGGCACTGGAGACACAGACTGCGACgaagacg
9F2 acatggcAAGACTACGAAGAAGACGACCTCGCGGACGCTTTGGCTTCTACCACAGAGGACGATGGCACTGGAGACACAGACTGCGACgaagacg
13F1 acatggcAAGACTACGAAGAAGACGACCTACGGACGCTTTGGCTTCTACCACAGAGGACGATGGCACTGGAGACACAGACTGCGACgaagacg
HGyV1-915 acatggcAAGACTACGAAGAAGACGACCTCGCGGACGCTTTGGCTTCTACCACAGAGGACGATGGCACTGGAGACACAGACTGCGACgaagacg
HGyV1-CL33 acatggcAAGACTACGAAGAAGACGACCTCGCGGACGCTTTGGCTTCTACCACAGAGGACGATGGCACTGGAGACACAGACTGCGACgaagacg
AGV2 acatggcAAGATTACGACGACGACGACCTCGTGGACGCTTTGGCTTCTACCACAGAGGACGATGGCACTGGAGACACAGACTGCGACgaagacg
GyV3 acatggcTAGA---CGATTTCCGCCGCACAGAGGAAAGTTCGGATACTACAGACGGGGAAGATGGCACTGGAGACACAGACTGCGACgaagacg
******** **** **** ** *** *** *** ** *** *********

Figure. Alignment of partial sequences of human gyroviruses (HGyVs) from healthy blood donors, France. Point mutation corresponding to a nonsynonymous substitution is in **boldface** (13F1 isolate). Reference sequences and GenBank accession nos.: HGyV1-915, FR823283; HGyV1-CL33, JQ308212; avian gyrovirus (AGV) 2, JQ690763; gyrovirus (GyV) 3, JQ308210). Bar above sequences indicates location of the HGyVsPBp probe; lowercase letters indicate 5'/3' ends of HGyVsPBs/HGyVsPBr real-time primers; asterisks (*) indicate conserved positions.
regions would be conserved differ-
ently must be considered.

It is probable that subsequent
sequences of HGyVs remain to be
identified in human blood. A recent
study reported the characterization
of a highly divergent GyV sequence
(GyV4) in human fecal samples and
chicken meat (7); as with avian GyV2
and GyV3, further research is needed
to determine whether this variant rep-
licates in the human body or is solely
ingested in food and passively excret-
ed. A better knowledge of the genetic
diversity of these newly discovered
viruses will enable development of
improved molecular detection sys-
tems and their subsequent use in epi-
demiologic studies involving diverse
human cohorts.

The potential clinical importance
of HGyVs remains to be clarified. Al-
though infection with CAV in birds
is frequently associated with clini-
cal signs and disease, the presence of
HGyVs in immunocompromised or
immunocompetent humans does not
appear to be correlated with visible
symptoms. Further studies of the natu-
ral history and distribution of HGyVs
in human hosts are needed.

Philippe Biagini,
Sandra Bédarida,
Mhammed Touinssi,
Vital Galicher, and
Philippe de Micco.

Author affiliation: Unités Mixtes de Recherche Centre National de la Recherche Scientifique 7268 Equipe "Emergence et coïvolution virale," Marseille, France

DOI: http://dx.doi.org/10.3201/eid1906.130228

References

1. Schat KA. Chicken anemia virus. Curr Top Microbiol Immunol. 2009;331:151-83. http://dx.doi.org/10.1007/978-3-540-70972-5_10

2. Sauvage V, Cheval J, Foulongne V, Ar Gouilh M, Pariente K, Manuguerra JC, et al. Identification of the first human gyrovirus, a virus related to chicken anemia virus. J Virol. 2011;85:7948-50. http://dx.doi.org/10.1128/JVI.00639-11

3. Rijsewijk FA, dos Santos HF, Teixeira TF, Cibulski SP, Varela AP, Dezen D, et al. Discovery of a genome of a distant relative of chicken anemia virus reveals a new member of the genus *Gyrovirus*. *Arch Virol*. 2011;156:1097-100. http://dx.doi.org/10.1007/s00705-011-0971-6

4. Phan TG, Li L, O'Ryan MG, Cortes H, Mamani N, Bonkoungou IJ, et al. A third gyrovirus species in human faeces. J Gen Virol. 2012;93:1356-61. http://dx.doi.org/10.1099/vir.0.041731-0

5. Maggi F, Macera L, Focosi D, Vatteroni ML, Boggi U, Antonelli G, et al. Human gyrovirus DNA in human blood, Italy. Emerg Infect Dis. 2012;18:956-9. http://dx.doi.org/10.3201/eid1806.120179

6. Biagini P, Dussol B, Touinssi M, Brunet P, Picard C, Moal V, et al. Human parvovirus 4 in kidney transplant patients, France. Emerg Infect Dis. 2008;14:1811-2. http://dx.doi.org/10.3201/eid1411.080862

7. Chu DK, Poon LL, Chiu SS, Chan KH, Ng EM, Bauer I, et al. Characterization of a novel gyrovirus in human stool and chicken meat. J Clin Virol. 2012;55:209-13. http://dx.doi.org/10.1016/j.jcv.2012.07.001

Address for correspondence: Philippe Biagini, UMR CNRS 7268 Equipe "Emergence et coïvolution virale," Etablissement Français du Sang Alpes-Méditerranée et Aix-Marseille Université, 27 Boulevard Jean Moulin, 13005 Marseille, France; email: philippe.biagini@efs.sante.fr

Vibrio cholerae
O1 Isolate with
Novel Genetic
Background,
Thailand–Myanmar

To the Editor: *Vibrio cholerae*
O1, a causative agent of cholera, was
classified into 2 biotypes, classical and
El Tor (I). However, accumulating
evidence suggests that atypical El Tor
*V. cholerae*, which possesses traits of
both classical and El Tor biotypes, has
replaced the seventh pandemic pro-
totypic El Tor *V. cholerae* worldwide
in recent years. Cholera outbreaks

in Thailand during 2007–2010 were
caused by atypical El Tor isolates car-
rying the classical type cholera toxin
gene (2). Epidemiologic surveys in a
Thailand–Myanmar border area during
2008–2012 yielded more than 500 iso-
lates of *V. cholerae* O1. We identified
an isolate that possessed the typical El
Tor type cholera toxin gene (genotype
3) and designated it MS6 (later as-
signed strain number DMST28216). It
does not belong to either the seventh
pandemic prototypic biotype identified
in 1961 or the group of atypical El Tor
strains found during 1991–present (3).

MS6 was isolated from stool
samples from a 26-year-old woman
(migrant worker) from Myanmar who
had been admitted to Mae Sot General
Hospital in Tak Province, Thailand,
for 3 days with vomiting, watery diar-
rhea, nausea, fever, and headache. The
illness was considered mild to mod-
erate. Acute gastroenteritis was diag-
nosed on the basis of the symptoms
and laboratory results. The key viru-
lence factors of *V. cholerae* O1 include
cholera toxin (CTX), which is respon-
sible for profuse watery diarrhea, and
a pilus colonization factor known as
toxin-coregulated pilus (TCP). The
virulence-related genes (*ctxAB* and
*tcpA*) and the phage repressor gene
(*rstR*) of MS6 had identical sequences
to those of the seventh pandemic pro-
totypic El Tor *V. cholerae* O1 N16961
strain. The isolate was found to be
positive for enteric bacteria in the
Voges-Proskauer test and resistant to
polymyxin B (50 units). We further
investigated 2 gene clusters, *Vibrio*
seventh pandemic island I (VSP-I)
and II (VSP-II), associated with the
seventh pandemic strains and absent
in classical and pre–seventh pandemic
strains (4–7). The common genes on
the VSP-I island in N16961, including
VC0175, VC0178, VC0180, VC0181,
and VC0183, were detected by PCR
(8) in MS6 but were lacking in VSP-
II; 26.9 kb of VSP-II was originally
found in N16961. Moreover, PCR
analysis showed that the isolate did not