Molecular detection and characterisation of herpesviruses in asymptomatic Russian sturgeon (Acipenser gueldenstaedtii) from European aquaculture

Fig. 1.

Fig. 2.

Fig. 3.

Assessments of percentage sequence similarities according to GenBank for PCR products of amplification of a disease aetiological agent isolated from different fishes

Fish	Acipenser ruthenus	AciHV-3	Remarks
Common carp	96%	99%
Tench	97%	99%
Crucian carp 1	97%	99%	AciHV-3-like
Crucian carp 2	95%	96%
Sturgeon 1	97%	99%
Sturgeon 2	90%	92%	Suspicious
Sturgeon 3	96%	98%
Sturgeon 4	97%	99%
Sturgeon 5	97%	99%	AciHV-3-like
Sturgeon 6	96%	98%
Sturgeon 7	97%	99%
Sturgeon 8	97%	99%
Average value	96%	98%
Median value	97%	99%

Primer pairs used in this study for DNA and RNA extraction from Russian sturgeon in a mortality event and from cohabited tench, Nile tilapia, crucian carp and common carp

Name	Sequence	Reference
Stör-HV-1F	5′-CGG AAT TCT AGA YTT YGC IWS IYT ITA YCC-3′	(9)
Stör-HV-1R	5′-CCC GAA TTC AGA TCT CIG TRT CIC CRT A-3′
Stör-HV-Fn	5′-GCA GCA GAC TAC GTG GTG TAC G-3′	(9)
Stör-HV-Rn	5′-AGT TGG RGC AGA TCT TCA TTT CGG-3′
Terminase-AciHV-1 F	5′-ACC TCG TGT TGA TCG-3′	(15)
Terminase-AciHV-1 R	5′-TCA AAA CTT CCG GGT-3′
Term F2	5′-GCM MGR GGA CAG AWC CCM G-3′	(15)
Termsal-3 Rdeg	5′-GGT GCA CAC RCC MAD IGA CG-3′
TermExon 1F	5′-CAG GTB GAR CTM ATG MGR GGG TTT TT-3′	personal
TermExon 1R	5′-CAT RAT KKY DGT YTT VCC RCA YTG TC-3′	communication

Preparation of master mixes for first-round PCRs using a 5 μL template to amplify DNA of an aetiological agent infecting Russian sturgeon lethally

Components	volume × 1 (in μL)
Diethyl pyrocarbonate–treated water	11.7
5× Green GoTaq Flexi Puffer (Promega)	5
MgCl2 25 mM (Promega)	2.5
dNTPs 10 mM (Thermo Fisher)	0.25
GoTaq DNA polymerase (Promega)	0.125
Primer premix (1:1) 400 nmol	0.5

RNA and DNA extraction kit concentration of DNA from an aetiological agent infecting Russian sturgeon lethally

Sturgeon sample numbers	RNA extraction kit double-stranded DNA in ng/μL*	DNA extraction kit
388	554.90	893.05
544	150.65	1220.65
577	889.55	889.05
608	240.95	2051.75
Average	459.0125	1263.625

Positive sturgeon herpesvirus PCRs in different fish samples

Species	Positive cases/total	Rate (%)	Remarks
Sturgeon	105/120	87.5	Infected
Tench	2/5	40	Infected
Crucian carp	4/6	66.6	Infected
Common carp	2/6	33.3	Suspicious but positive
Tilapia	0/4	-	Negative

Investigation of sturgeon organ tropism using PCR for sturgeon herpesvirus

Organ	Sampling
	1 (n = 359)	2 (n = 360)	3 (n = 361)	4 (n = 362)	5 (n = 363)
Heart	+	+	+	-	+
Liver	+	+	+	-	+
Spleen	-	+	+	-	+
Kidney	-	-	+	-	+
Pancreas	-	+	+	-	+
Swim bladder	-	+	+	-	+