Saturday, June 25, 2016

PLoS NTD: Isolation of Infective Zika Virus from Urine and Saliva of Patients in Brazil


Until a few months ago sexual transmission of the Zika virus was considered to be a rare - almost theoretical - possibility. 

Prior to that there had been only one documented case (see Probable Non–Vector-borne Transmission of Zika Virus, Colorado, USA) involving an infected researcher in Africa who returned to the United States in 2008 and passed the virus (presumably via sexual contact) to his wife.

But over the past five months the United States has reported 11 cases of sexual transmission of the virus, and similar reports have been received from around the world.

In regions with a competent mosquito vector - which is still the most common source of Zika infection - parsing out sexually transmitted infections from the rest is nearly impossible.   

While initially believed to be a minor component in the chain of transmission, Zika is gaining a reputation as being an STD (see CDC : Sexual Transmission Of Zika May Be More Common Than Previously Believed), and the CDC has released comprehensive guidance on the prevention of sexual transmission.

In February, researchers in Brazil reported finding live Zika virus and Zika RNA in both saliva and urine (see FIOCRUZ: Statement On Detection Of Zika Virus In Saliva and Urine).

Another study, appearing in Eurosurveillance: Prolonged Shedding Of Zika Virus RNA In Saliva - Italy in early March, also reported on the detection of both live virus and viral RNA, and suggested their findings  pose questions on the potential risk of human-to-human transmission of the virus through saliva.

This oral transmission scenario was revisited again earlier this month in NEJM: Possibility Of Oral Transmission Of Zika Virus, where French researchers reported on a sexually transmitted case where oral transmission could not be `ruled out'.  

While plausible, the oral route of Zika transmission remains less than proven.

Yesterday PLoS Neglected Tropical Diseases published the study from researchers at the FIOCRUZ (Fundação Oswaldo Cruz) Institute that first raised serious questions over the possibility of oral transmission of the virus last February.

Research Article

Isolation of Infective Zika Virus from Urine and Saliva of Patients in Brazil

Myrna C. Bonaldo , Ieda P. Ribeiro , Noemia S. Lima , Alexandre A. C. dos Santos , Lidiane S. R. Menezes , Stephanie O. D. da Cruz , Iasmim S. de Mello , Nathália D. Furtado , Elaine E. de Moura , Luana Damasceno , Kely A. B. da Silva , Marcia G. de Castro , Alexandra L. Gerber , Luiz G. P. de Almeida , Ricardo Lourenço-de-Oliveira , Ana Tereza R. Vasconcelos , Patrícia Brasil

Published: June 24, 2016



Zika virus (ZIKV) is an emergent threat provoking a worldwide explosive outbreak. Since January 2015, 41 countries reported autochthonous cases. In Brazil, an increase in Guillain-Barré syndrome and microcephaly cases was linked to ZIKV infections. A recent report describing low experimental transmission efficiency of its main putative vector, Ae. aegypti, in conjunction with apparent sexual transmission notifications, prompted the investigation of other potential sources of viral dissemination. Urine and saliva have been previously established as useful tools in ZIKV diagnosis. Here, we described the presence and isolation of infectious ZIKV particles from saliva and urine of acute phase patients in the Rio de Janeiro state, Brazil.

Methodology/Principal Findings

Nine urine and five saliva samples from nine patients from Rio de Janeiro presenting rash and other typical Zika acute phase symptoms were inoculated in Vero cell culture and submitted to specific ZIKV RNA detection and quantification through, respectively, NAT-Zika, RT-PCR and RT-qPCR. Two ZIKV isolates were achieved, one from urine and one from saliva specimens. ZIKV nucleic acid was identified by all methods in four patients. Whenever both urine and saliva samples were available from the same patient, urine viral loads were higher, corroborating the general sense that it is a better source for ZIKV molecular diagnostic. In spite of this, from the two isolated strains, each from one patient, only one derived from urine, suggesting that other factors, like the acidic nature of this fluid, might interfere with virion infectivity. The complete genome of both ZIKV isolates was obtained. Phylogenetic analysis revealed similarity with strains previously isolated during the South America outbreak.


The detection of infectious ZIKV particles in urine and saliva of patients during the acute phase may represent a critical factor in the spread of virus. The epidemiological relevance of this finding, regarding the contribution of alternative non-vectorial ZIKV transmission routes, needs further investigation.

Author Summary

The American continent has recently been the scene of a devastating epidemic of Zika virus and its severe manifestations, such as microcephaly in newborns and Guillain-Barré Syndrome. Zika virus, first detected in 1947 in Africa, only from 2007 started provoking outbreaks. Zika, dengue and chikungunya viruses are primarily transmitted by Aedes mosquitoes. Dengue has been endemic in Brazil for almost 30 years, and the country is largely infested by its main vector, Aedes aegypti. Chikungunya virus entered the country in late 2014 and Zika presence was confirmed eight months later.
Nevertheless, Zika notifications multiplied and spread across the country with unprecedented speed, raising the possibility of other transmission routes. This hypothesis was strengthened by some recent reports of Zika sexual transmission in Ae. aegypti-free areas and by the description of a low transmission efficiency to Zika virus in local Ae. aegypti. We found Zika active particles in both urine and saliva of acute phase patients, and a finding that was promptly announced by Fiocruz via Press Conference on February 5, 2016. In this work, we bring up the potential alternative person-to-person infection routes beyond the vectorial transmission, that might have epidemiological relevance.
(Continue . . . )

With a relatively small cohort of sexually transmitted cases available for study, trying to parse out exactly what kind of intimate contact led to the infection of their partner is a difficult task.

But as the number of cases rise in areas without a competent mosquito vector, coupled with the increased adoption of  Zika `safe sex' practices, the sampling size may eventually allow for a more definitive answer.

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