Dados do Trabalho

Título

Usefulness of saliva samples for hepatitis C virus genotyping using nucleotide sequencing

Introdução

It is estimated that 58 million people are chronically infected with hepatitis C virus (HCV). In May 2016, the World Health Assembly proposed the elimination of viral hepatitis as a public health threat by 2030 which includes an increase in diagnosis access by using alternative specimens as saliva since it has its advantages, such as minor risk of transmission, less invasive and the option of self-collection. Also, assessing viral genetic diversity by sequencing at other fluids beyond gold standard blood samples can facilitate HCV genomic surveillance, which enables detection and study of RAS (resistance-associated substitutions). Genotyping can also be important to failure-patients given that 1 and 3 genotypes are more associated with cirrhosis and other negative outcomes.

Objetivo (s)

Evaluate saliva as an alternative specimen for HCV genotyping by conventional sequencing.

Material e Métodos

A total of 36 individuals referred to Viral Hepatitis Ambulatory (AHV/IOC) were recruited for biological collection, being 21 with detectable HCV RNA at RT-qPCR in serum and 15 healthy subjects (HCV RNA negative). Viral RNA was extracted by commercial kit where sample volume was twofold increased for saliva samples. HCV NS5B gene was amplified by two-round PCR (~370 nucleotides). PCR products were purified, quantified, and submitted to Sanger sequencing.

Resultados e Conclusão

RESULTS: The thirty-six patients comprised 63,88% (23/36) by women and 36.11% (13/36) by men. The mean age was 56± 8,95 years. Mean viral load in serum was 5.878 ± 0.645 log10 IU/mL. HCV RNA was not detected in saliva from negative individuals (0/15), and it was detected in 7/21 (33.3%) saliva samples from the HCV group. The main HCV genotype at serum was 1 (71.42%, 15/21). After Sanger sequencing, it was possible to obtain three good HCV nucleotide sequences in saliva. Paired samples presented the same genotypes and similar sequences in both specimens 1a: 2 of 3 sets; 1b: 1 of 3 sets. CONCLUSIONS: It was possible to detect HCV RNA in saliva samples although the sensitivity was low. In addition, nucleotide sequencing was efficient to demonstrate HCV genotypes similar in serum and saliva samples indicating the potential of this specimen for molecular epidemiological studies.

Palavras-chave

HCV, genotypes, saliva, sequencing

Agradecimentos

CNPq, FAPERJ