HCV genotype distribution and determination of other variables related to the
virus by assessing a clinical laboratory results database
CHIANCA, C.F.1; SANTA RITA, T.H.1; COSTA, P.G.G.2; VIVAS, W.L.P.3; VELASCO, L.F.R2; ABDALLA, L.F.2; COSTA, S.S.S.2; BARRA, G.B.2;
1Universidade
de Brasília, Brasilia, Brazil; 2Laboratório Sabin de Análises Clínicas, Brasilia, Brazil; 3Universidade Tiradentes, Aracaju, Brazil
Table 1: Distribution of genotypes in relation to gender and age range
Abstract
Introduction
In 2011, FDA announced a recall of an in vitro nucleic acid amplification test for the quantitation of HCV RNA that has been shown to under-quantitate a subset of genotype 4 in patient
specimens by approximately 1.0-1.5 log 10 in the absence of any sequence mismatches. The distribution of these test include your country. As the HCV genotype has significant
geographic variation, this recall encorage us to determine the prevalence of HCV genotypes and others variables releted to the virus by assessing our clinical laboratory results database,
and evaluate the possible impact of this under-quantitation in our region.
Methods
Through retrospective analysis of our HCV genotyping database, we assessed the samples results between January 2005 and September 2011. 480 samples were analyzed, 394 (82%)
genotypes were determined and 86 (17.9%) had negative results (no genotype detected). Of these, 280 (58,2%) perform quantitation of HCV RNA in the same day. The genotype
prevalences and RNA quantification were identified and presented by gender and age group <50 years (n=162) and ≥50 years (n=118). HCV genotyping was performed by three different
techniques in the analyzed period, PCR-RFLP, reverse hibridization, and RT-qPCR, they were also compared.
Results
The obeserved genotype distribution was HCV-1 288 (73%), HCV-3 91 (23%), HCV-2 13 (3.2%), and HCV-4 2 (0.5%). The genotypes HCV-6 and HCV-5 were not found. The three techniques
show related capabilities, since they result in similar genotypic distributions frequency (p=0.98). The average viral load was 6.11 log10 copies/ml and did not differ with genotype (p=0.48),
or gender of the patient (p=0.54). By separating the sample into age groups <50 and ≥50 years the average viral load was 6.13 and 6.41 log10 copies/ml (P=0.032) respectively.
Conclusion
In this study we noted that the distribution of HCV in our samples is HCV-1, HCV-3, HCV-2 and HCV-4. The genotype 4 is rare in our region, as was found in only two individuals, or 0.5% of
samples in six and half years. Thus, the impact of under-quantitation in our region would be minimal. When comparing the different techniques used for molecular genotyping showed
that all three techniques (RT-qPCR real-time Reverse Hybridization and PCR-RFLP) have similar capabilities to determine the genotypes. With respect to viral load, no difference was
observed between genotypes, or in genotypes according to gender, nor in the genotypes according to age. However, the group aged ≥50 had a mean viral load significantly greater than
<50.
GENOTYPE
AGE RANGE
1
2
3
4
TOTAL
10 a 19
20 a 29
30 a 39
40 a 49
50 a 59
60 a 69
70 a 79
80 a 89
1 (0,7%)
6 (3,9%)
19 (12,4%)
35 (22,9%)
23 (15,0%)
24 (15,7%)
3 (2,0%)
2 (1,3%)
0 (0,0%)
0 (0,0%)
2 (1,3%)
2 (1,3%)
0 (0,0%)
1 (0,7%)
2 (1,3%)
0 (0,0%)
0 (0,0%)
3 (2,0%)
6 (3,9%)
10 (6,5%)
9 (5,9%)
3 (2,0%)
2 (1,3%)
0 (0,0%)
0 (0,0%)
0 (0,0%)
0 (0,0%)
0 (0,0%)
0 (0,0%)
0 (0,0%)
0 (0,0%)
0 (0,0%)
1 (0,7%)
9 (5,9%)
27 (17,6%)
47 (30,7%)
32 (20,9%)
28 (18,3%)
7 (4,6%)
2 (1,3%)
TOTAL
113 (73,9%)
7 (4,6%)
33 (21,6%)
0 (0,0%)
153 (100,0%)
1
2
3
4
TOTAL
10 a 19
20 a 29
30 a 39
0 (0,0%)
9 (3,7%)
23 (9,5%)
0 (0,0%)
0 (0,0%)
0 (0,0%)
0 (0,0%)
2 (0,8%)
10 (4,1%)
0 (0,0%)
0 (0,0%)
1 (0,4%)
0 (0,0%)
11 (4,6%)
34 (14,1%)
40 a 49
59 (24,5%)
2 (0,8%)
26 (10,8%)
1 (0,4%)
88 (36,5%)
treatment time. In 2011, FDA announced a recall of an in vitro nucleic acid amplification test for the quantitation of
50 a 59
60 a 69
70 a 79
80 a 89
69 (28,6%)
10 (4,1%)
4 (1,7%)
1 (0,4%)
4 (1,7%)
0 (0,0%)
0 (0,0%)
0 (0,0%)
18 (7,5%)
2 (0,8%)
0 (0,0%)
0 (0,0%)
0 (0,0%)
0 (0,0%)
0 (0,0%)
0 (0,0%)
91 (37,8%)
12 (5,0%)
4 (1,7%)
1 (0,4%)
HCV RNA that has been shown to under-quantitate a subset of genotype 4 in patient specimens by approximately
TOTAL
175 (72,6%)
6 (2,5%)
58 (24,1%)
2 (0,8%)
241 (100,0%)
AGE RANGE
Background
Hepatitis C is a silent disease, often discovered by routine screening, blood donation, among others. Brazil has a
high rate of morbidity and mortality and it is necessary to perform data collection on this situation. The
MALE
determination of the genotype and quantification of viral load is important in clinical practice to guide patient
1.0-1.5 log 10 in the absence of any sequence mismatches. The distribution of known genotypes and subtypes of
Reverse hibridization
60,0%
3 are more common in Europe, Japan and the United States, genotype 4 in Central Africa, Egypt and Middle East,
50,0%
Relative Frequency (%)
HCV shows significant geographic variation in the frequency with which they are observed. The genotypes 1,2 and
genotype 5 in South Africa and genotype 6 in Asia. In some regions in Brazil, the genotypes of hepatitis C virus has
not been widely studied, for example, the Federal District, Brazil.
Objetive
PCR-RFLP
RT-qPCR
Mean viral load Log10 (copies/ml)
FEMALE
40,0%
30,0%
20,0%
10,0%
0,0%
Determine the prevalence of HCV genotypes and others variables releted to the virus by assessing our clinical
1
2
3
4
GENOTYPE
GENOTYPE
laboratory results database, and evaluate the possible impact of this under-quantitation in our region.
Figure 2: Analysis of the mean viral load in log10 (copy / ml)
according to the genotypes.
Graph 1. Comparison of Techniques with genotypes.
Viral load Log10 (copies/ml)
Through retrospective analysis of our HCV genotyping database, we assessed the samples results between January
Mean viral load Log10 (copies/ml)
Methods
2005 and September 2011. 480 samples were analyzed, 394 (82%) genotypes were determined and 86 (17.9%) had
negative results (no genotype detected). Of these, 280 (58,2%) perform quantitation of HCV RNA in the same day.
The genotype prevalences and RNA quantification were identified and presented by gender and age group <50
years (n=162) and ≥
50 years (n=118). HCV genotyping was performed by three different techniques in the analyzed
period, PCR-RFLP, reverse hibridization, and RT-qPCR, they were also compared. Data from this study were
tabulated on an Excel ®. The graphs and tables of descriptive analysis will be generated with the aid of the Excel ®,
MALE
Prism ® and SPSS ®.
FEMALE
GROUP < 50
Figure 3: Analysis of the mean viral load in log10 (copy / ml)
according to the genre.
Results
GROUP ≥ 50
Figure 4: Analysis of the mean viral load in log10 (copy / ml)
with the groups <50 and ≥ 50 years.
Conclusion
The obeserved genotype distribution was HCV-1 288 (73%), HCV-3 91 (23%), HCV-2 13 (3.2%), and HCV-4 2 (0.5%).
The genotypes HCV-6 and HCV-5 were not found. In the present study, there was predominance of male gender
In this study we noted that the distribution of HCV in our samples is HCV-1, HCV-3, HCV-2 and HCV-4. The
and age group of 40-49 years.The three techniques show related capabilities, since they result in similar genotypic
genotype 4 is rare in our region, as was found in only two individuals, or 0.5% of samples in six and half years.
distributions frequency (p=0.98). The average viral load was 6.11 log10 copies/ml and did not differ with genotype
Thus, the impact of under-quantitation in our region would be minimal. When comparing the different techniques
(p=0.48), or gender of the patient (p=0.54). By separating the sample into age groups <50 and ≥50 years the
used for molecular genotyping showed that all three techniques (RT-qPCR real-time Reverse Hybridization and
average viral load was 6.13 and 6.41 log10 copies/ml (P=0.032) respectively.
PCR-RFLP) have similar capabilities to determine the genotypes. With respect to viral load, no difference was
observed between genotypes, or in genotypes according to gender, nor in the genotypes according to age.
However, the group aged ≥50 had a mean viral load significantly greater than <50.
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