Volume 5 - 2024 | https://doi.org/10.3389/fitd.2024.1489647 Introduction: Zika virus (ZIKV) is primarily transmitted through mosquito bites though may act as a sexually transmitted infection Men can shed the virus in their semen for extended periods with reports indicating shedding for up to six months after the initial infection This poses a concern for couples planning pregnancy whether through natural or assisted methods due to the risk of congenital Zika disease Human reproductive clinics typically perform serologic tests to screen for Zika infection we evaluated semen samples stored in a human reproduction clinic in Salvador using real-time quantitative reverse transcription polymerase chain reaction (RT-qPCR) technique Results: 36 sperm samples from 23 donors were analyzed including 5 with positive IgG serology for ZIKV and all negative for IgM all analyzed samples were negative by RT-qPCR ZIKV Discussion: Despite the limited number of samples this study raises further concern that the use of serology may be an unreliable surrogate method to predict the presence of ZIKV in semen Zika virus (ZIKV) is an RNA flavivirus with the first documented case of ZIKV infection reported in 1947 in the Zika Forest in Uganda. In 2013 an outbreak occurred in French Polynesia spreading widely throughout the world including South America (1) including testing of couples and semen donors It remains unknown whether immunoglobulin tests for ZIKV are sufficiently accurate to determine presence of prior infection and eligibility as a semen donor The present study seeks to evaluate whether serologic tests are reliable for screening semen donors for ZIKV compared with virus screening using RT-qPCR we selected samples from a sperm bank at the Instituto Valenciano de Infertilidade (IVI) which was the epicenter of the ZIKV outbreak in Brazil These samples were chosen from aliquots designated for discard These samples were collected between 2013 and 2019 a time window that encompasses both the beginning of the epidemic period of ZIKV and the years following it in Brazil We analyzed socio-demographic data and serologic tests for ZIKV (IgM and IgG) performed at the time of each semen donation as part of standard screening Cryopreserved semen samples were then tested for the presence of ZIKV using RT-qPCR This study received approval from the ethics and research committee of the Faculdade de Medicina da Universidade Federal da Bahia For the molecular diagnosis of ZIKV infection Viral RNA was initially extracted using a modified two-step protocol that involved TRIzol Reagent-based extraction followed by a magnetic bead-based technology for further purification we combined 200 µL of sperm samples with 800 µL of TRIzol Reagent (Thermo Fisher Scientific 15596026) and 160 µL of chloroform (Sigma-Aldrich then centrifuged the mixture for 15 minutes at 12,000xg at 4°C The resulting aqueous phase was subjected to additional purification using magnetic bead-based technology to ensure high-quality RNA extraction This further purification was conducted using the MagMAX Viral/Pathogen II (MVP II) Nucleic Acid Isolation Kit (Thermo Fisher Scientific A48383R) on the KingFisher Flex System (Thermo Fisher Scientific all according to the manufacturer’s instructions ZIKV detection was carried out using the ZDC Molecular Kit (Instituto de Biologia Molecular do Paraná 80142170032) on a 7500 Real-Time PCR System (Applied Biosystems also following the manufacturer’s instructions Cq values to determine sensitivity for ZIKV detection in sperm samples RT-qPCR semen samples and IgG serology results of the participants candidates for assisted fertilization undergo serological testing RT-qPCR testing of the collected semen samples is not commonly performed serological testing may result in false positive testing of ongoing ZIKV infection particularly in areas with other endemic arboviruses (Dengue in particular) that may have demonstrate cross-reactivity serologically the duration and viability of ZIKV in samples from various body fluids may not correlate with serological test results This test demonstrates excellent specificity and sensitivity for Zika virus diagnosis without the risk of cross-reactivity with other arboviruses as seen in traditional serology utilized in the current study it is a labor-intensive test requiring several days with a limited number of samples able to be tested simultaneously further studies investigating the correlation between virus persistence in semen and IgM titers are required to evaluate the utility of IgM as a biomarker of ZIKV in semen including a small sample size of semen samples analyzed which were obtained through convenience sampling positive IgM or IgG identified in our study by standard serology may be false positive for ZIKV given cross-reactivity with DENV and may limit interpretation of discordant serologic positive and negative RT-qPCR results With a new outbreak of Zika infection in the future a prospective study may determine the accuracy of different diagnostic tests (serologic PRNT and MAC-ELISA and nucleic acid testing) over a specified timeline during and after infection This approach would aid in the development of diagnostic algorithms and evaluate their real-world reliability and cost-effectiveness for individuals of reproductive age and assisted reproduction clinics Several unresolved questions linger: Is it necessary to test semen samples using the RT-qPCR technique Would serological testing alone be sufficient for a couple in which one partner has traveled to the ZIKV endemic zone Does a positive RT-qPCR in the semen correspond to viable viruses with the potential for infection An approach based on available data may include condom use for the first 30 days (about 4 and a half weeks) following possible ZIKV exposure the next recommended test would be the ZIKV IgM MAC-ELISA which can indicate a probable recent infection it would be considered safe for spontaneous or assisted contraception where infection can be transmitted by mosquitoes at any time routine precautions against mosquito bites must be maintained The raw data supporting the conclusions of this article will be made available by the authors The studies involving humans were approved by Faculdade de Medicina da Universidade Federal da Bahia The studies were conducted in accordance with the local legislation and institutional requirements The participants provided their written informed consent to participate in this study The author(s) declare financial support was received for the research This research was funded by the Coordenação de Aperfeiçoamento de Pessoal de Nível Superior—Brasil (CAPES) [Finance Code 001] Conselho Nacional de Desenvolvimento Cientfico e Tecnológico [439967/2016-3] Programa de Excelência em Pesquisa [420765/2017-4] and Financiadora de Estudos e Projetos [MCTIC/FINEP/FNDCT 01/2016 ZIKA—0416006000] The funding organizations had no role in the study design The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest The author(s) declared that they were an editorial board member of Frontiers This had no impact on the peer review process and the final decision All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations Any product that may be evaluated in this article or claim that may be made by its manufacturer is not guaranteed or endorsed by the publisher Persistence and clinical relevance of Zika virus in the male genital tract Differences in prevalence of symptomatic zika virus infection In: Zika virus disease - Interim case definition (2016) Google Scholar Zika virus: High infectious viral load in semen The re-emergence of Zika in Brazil in 2020: A case of Guillain Barré Syndrome during the low season for arboviral infections Zika virus in the Americas: Early epidemiological and genetic findings Characterizing the pattern of anomalies in congenital zika syndrome for pediatric clinicians doi: 10.1001/jamapediatrics.2016.3982 Potential sexual transmission of zika virus Suspected female-to-male sexual transmission of Zika virus — New York City Male-to-male sexual transmission of Zika virus — Texas Google Scholar Persistent detection of Zika virus RNA in semen for six months after symptom onset in a traveller returning from Haiti to Italy doi: 10.2807/1560-7917.ES.2016.21.32.30314 Effect of acute Zika virus infection on sperm and virus clearance in body fluids: a prospective observational study Persistence and intra-host genetic evolution of Zika virus infection in symptomatic adults: A special view in the male reproductive system Estimate of the reproduction number of the 2015 Zika virus outbreak in Barranquilla and estimation of the relative role of sexual transmission Prevention and control of zika as a mosquito-borne and sexually transmitted disease: a mathematical modeling analysis Sexual transmission of Zika virus and other flaviviruses: A living systematic review WHO guidelines for the prevention of sexual transmission of Zika virus Google Scholar Update: interim guidance for preconception counseling and prevention of sexual transmission of zika virus for men with possible Zika virus exposure — United States An update on the aspects of Zika virus infection on male reproductive system Google Scholar Zika virus shedding in semen of symptomatic infected men Accuracy of the Zika IgM antibody capture enzyme-linked immunosorbent assay from the centers for disease control and prevention (CDC Zika MAC-ELISA) for diagnosis of Zika virus infection Persistence of Zika virus in body fluids — final report Santos LA and Khouri R (2025) Screening semen samples for Zika virus infection: role for serologic and RT-PCR testing Received: 02 September 2024; Accepted: 23 December 2024;Published: 29 January 2025 Copyright © 2025 Ferraz, Akrami, de Moraes, Gusmão, Coelho, Freitas, Barral-Netto, Barral, Boaventura, Santos and Khouri. This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY) distribution or reproduction in other forums is permitted provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited in accordance with accepted academic practice distribution or reproduction is permitted which does not comply with these terms *Correspondence: Ricardo Khouri, cmljYXJkby5raG91cmlAZmlvY3J1ei5icg== Disclaimer: All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations Any product that may be evaluated in this article or claim that may be made by its manufacturer is not guaranteed or endorsed by the publisher 94% of researchers rate our articles as excellent or goodLearn more about the work of our research integrity team to safeguard the quality of each article we publish Posted in: Comics | Tagged: That the recent CCXP 2018 comic book convention in Brazil saw the release of the DC Comics crossover with Monica's Gang characters in a variety of forms Monica's Gang is America's best selling comic… it just happens to mostly sell in South America Created by Mauricio de Sousa and based on a newspaper comic strip from 1959 over the years the series has gained a large audience with new characters constantly being added to the lineup spinning off. Jimmy Five (Cebolinha) and Monica into her own comic book The stories revolve around the adventures of Monica and her many friends in a fictional neighborhood known as "Lemon Tree" inspired by the neighborhood of Cambuí in Campinas and the city of Mogi das Cruzes Published in the US as Monica And Her Friends she is now getting a bunch of Superfriends As well as being published in the usual cartoony Monica's Gang style With special attention towards Capitão Pitoco a character already created as a parody of Superman within the Monica's Gang world drawn closer to the DC Comics house style by artists who often draw for DC Comics anyway There seem no plans to publish this set of crossover comics in the USA But they took pride of place at the show Could the grey market turn these volumes into collector items in the USA Keep an eye on eBay and Amazon Marketplace… The dates displayed for an article provide information on when various publication milestones were reached at the journal that has published the article activities on preceding journals at which the article was previously under consideration are not shown (for instance submission All content on this site: Copyright © 2025 Elsevier B.V.