ABSTRACT
Many virus lysis/transport buffers used in molecular diagnostics, including the detection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) RNA, contain guanidine-based chaotropic salts, primarily guanidine hydrochloride (GuHCl) or guanidine isothiocyanate (GITC). Although the virucidal effects of GuHCl and GITC alone against some enveloped viruses have been established, standardized data on their optimum virucidal concentrations against SARS-CoV-2 and effects on viral RNA stability are scarce. Thus, we aimed to determine the optimum virucidal concentrations of GuHCl and GITC against SARS-CoV-2 compared to influenza A virus (IAV), another enveloped respiratory virus. We also evaluated the effectiveness of viral RNA stabilization at the determined optimum virucidal concentrations under high-temperature conditions (35°C) using virus-specific real-time reverse transcription polymerase chain reaction. Both viruses were potently inactivated by 1.0 M GITC and 2.5 M GuHCl, but the GuHCl concentration for efficient SARS-CoV-2 inactivation was slightly higher than that for IAV inactivation. GITC showed better viral RNA stability than GuHCl at the optimum virucidal concentrations. An increased concentration of GuHCl or GITC increased viral RNA degradation at 35°C. Our findings highlight the need to standardize GuHCl and GITC concentrations in virus lysis/transport buffers and the potential application of these guanidine-based salts alone as virus inactivation solutions in SARS-CoV-2 and IAV molecular diagnostics.
Subject(s)
Guanidine , Influenza A virus , RNA, Viral , SARS-CoV-2 , Specimen Handling , SARS-CoV-2/drug effects , SARS-CoV-2/genetics , Influenza A virus/drug effects , Influenza A virus/genetics , Guanidine/pharmacology , Guanidine/chemistry , RNA, Viral/genetics , Humans , Specimen Handling/methods , Genome, Viral , COVID-19/virology , COVID-19/diagnosis , Chlorocebus aethiops , Vero Cells , Virus Inactivation/drug effects , Animals , RNA Stability/drug effects , Containment of Biohazards , Guanidines/pharmacology , Guanidines/chemistry , Salts/pharmacology , Salts/chemistryABSTRACT
The continuous evolution of H5Nx highly pathogenic avian influenza viruses (HPAIVs) is a major concern for accurate diagnosis. We encountered some challenges in subtyping and sequencing a recently isolated H5N1 HPAIV strain using classical diagnostic methods. Oropharyngeal, conjunctival, and cloacal swabs collected from a dead white-tailed eagle (Haliaeetus albicilla albicilla) were screened via real-time RT-PCR targeting the influenza A virus matrix (M) gene, followed by virus isolation. The hemagglutination inhibition test was applied in order to subtype and antigenically characterize the isolate using anti-A/duck/Hong Kong/820/80 (H5N3) reference serum or anti-H5N1 cross-clade monoclonal antibodies (mAbs). Sequencing using previously reported universal primers was attempted in order to analyze the full-length hemagglutinin (HA) gene. Oropharyngeal and conjunctival samples were positive for the M gene, and high hemagglutination titers were detected in inoculated eggs. However, its hemagglutination activity was not inhibited by the reference serum or mAbs. The antiserum to a recently isolated H5N1 clade 2.3.4.4b strain inhibited our isolate but not older strains. A homologous sequence in the previously reported forward primer and HA2 region in our isolate led to partial HA gene amplification. Finally, next-generation sequencing confirmed the isolate as H5N1 clade 2.3.4.4b HPAIV, with genetic similarity to H5N1 strains circulating in Japan since November 2021.
Subject(s)
Influenza A Virus, H5N1 Subtype , Influenza A virus , Influenza in Birds , Animals , Hemagglutinin Glycoproteins, Influenza Virus/genetics , Hemagglutinins , Influenza A virus/genetics , Japan/epidemiology , Seasons , BirdsABSTRACT
Background: In 2009, Kenyatta National Hospital (KNH) integrated cervical cancer screening within HIV care using visual inspection with acetic acid (VIA) and Pap smear cytology. Objectives: We evaluated utilisation of cervical cancer screening and human papillomavirus (HPV) vaccination among women living with HIV (WLHIV) receiving HIV care at KNH. Method: From November 2019 to February 2020, WLHIV aged ≥ 14 years were invited to participate in a survey following receipt of routine HIV services. We assessed awareness of cervical cancer, uptake of cervical cancer screening, uptake of the HPV vaccine, and barriers to utilisation of these services. In a subset of survey participants, focus group discussions (FGDs) were also conducted to identify screening barriers. Results: Overall, 305 WLHIV participated in the survey. Median age was 36 years (interquartile range [IQR]: 28-43), 41% were married, and 38% completed secondary education. Most (90%) had HIV RNA < 1000 copies/mL. Awareness of cervical cancer was high (84%), although only 45% of WLHIV had screened for cervical cancer at the referral hospital and only 13% knew how to prevent high-risk HPV. No participants had received an HPV vaccination. Older age, higher education, and knowledge of the HPV vaccine were associated with higher likelihood of cervical cancer screening (P < 0.05). In FGDs, barriers to utilising the services included user fees, fear of the procedure impacting fertility, age and gender of the provider, and long waiting times. Conclusion: Despite integration with HIV services, the utilisation of cervical cancer screening was low among WLHIV and implementation barriers contributed to low utilisation.
