Influenza A viral loads in respiratory samples collected from patients infected with pandemic H1N1, seasonal H1N1 and H3N2 viruses

BACKGROUND: Nasopharyngeal aspirate (NPA), nasal swab (NS), and throat swab (TS) are common specimens used for diagnosis of respiratory virus infections based on the detection of viral genomes, viral antigens and viral isolation. However, there is no documented data regarding the type of specimen that yields the best result of viral detection. In this study, quantitative real time RT-PCR specific for M gene was used to determine influenza A viral loads present in NS, NPA and TS samples collected from patients infected with the 2009 pandemic H1N1, seasonal H1N1 and H3N2 viruses. Various copy numbers of RNA transcripts derived from recombinant plasmids containing complete M gene insert of each virus strain were assayed by RT-PCR. A standard curve for viral RNA quantification was constructed by plotting each Ct value against the log quantity of each standard RNA copy number. RESULTS: Copy numbers of M gene were obtained through the extrapolation of Ct values of the test samples against the corresponding standard curve. Among a total of 29 patients with severe influenza enrolled in this study (12 cases of the 2009 pandemic influenza, 5 cases of seasonal H1N1 and 12 cases of seasonal H3N2 virus), NPA was found to contain significantly highest amount of viral loads and followed in order by NS and TS specimen. Viral loads among patients infected with those viruses were comparable regarding type of specimen analyzed. CONCLUSION: Based on M gene copy numbers, we conclude that NPA is the best specimen for detection of influenza A viruses, and followed in order by NS and TS

Serological Response to the 2009 Pandemic Influenza A (H1N1) Virus for Disease Diagnosis and Estimating the Infection Rate in Thai Population

BACKGROUND: Individuals infected with the 2009 pandemic virus A(H1N1) developed serological response which can be measured by hemagglutination-inhibition (HI) and microneutralization (microNT) assays. METHODOLOGY/PRINCIPAL FINDINGS: MicroNT and HI assays for specific antibody to the 2009 pandemic virus were conducted in serum samples collected at the end of the first epidemic wave from various groups of Thai people: laboratory confirmed cases, blood donors and health care workers (HCW) in Bangkok and neighboring province, general population in the North and the South, as well as archival sera collected at pre- and post-vaccination from vaccinees who received influenza vaccine of the 2006 season. This study demonstrated that goose erythrocytes yielded comparable HI antibody titer as compared to turkey erythrocytes. In contrast to the standard protocol, our investigation found out the necessity to eliminate nonspecific inhibitor present in the test sera by receptor destroying enzyme (RDE) prior to performing microNT assay. The investigation in pre-pandemic serum samples showed that HI antibody was more specific to the 2009 pandemic virus than NT antibody. Based on data from pre-pandemic sera together with those from the laboratory confirmed cases, HI antibody titers ≥ 40 for adults and ≥ 20 for children could be used as the cut-off level to differentiate between the individuals with or without past infection by the 2009 pandemic virus. CONCLUSIONS/SIGNIFICANCE: Based on the cut-off criteria, the infection rates of 7 and 12.8% were estimated in blood donors and HCW, respectively after the first wave of the 2009 influenza pandemic. Among general population, the infection rate of 58.6% was found in children versus 3.1% in adults

Lack of transmission among healthcare workers in contact with a case of Middle East respiratory syndrome coronavirus infection in Thailand

Abstract Introduction A hospital-associated outbreak of Middle East Respiratory Syndrome Coronavirus (MERS-CoV) was reported. We aimed to assess the effectiveness of infection control measures among healthcare workers (HCWs) who were exposed to a MERS patient and/or his body fluids in our institute. Methods A descriptive study was conducted among HCWs who worked with a MERS patient in Bamrasnaradura Infectious Diseases Institute, Thailand, between 18 June and 3 July 2015. Contacts were defined as HCWs who worked in the patient’s room or with the patient’s body fluids. Serum samples from all contacts were collected within 14 days of last contact and one month later. Paired sera were tested for detection of MERS‐CoV antibodies by using an indirect ELISA. Results Thirty-eight (88.4 %) of 43 identified contacts consented to enroll. The mean (SD) age was 38.1 (11.1) years, and 79 % were females. The median (IQR) cumulative duration of work of HCWs in the patient’s room was 35 (20–165) minutes. The median (IQR) cumulative duration of work of HCWs with the patient’s blood or body fluids in laboratory was 67.5 (43.7–117.5) minutes. All contacts reported 100 % compliance with hand hygiene, using N95 respirator, performing respirator fit test, wearing gown, gloves, eye protection, and cap during their entire working period. All serum specimens of contacts tested for MERS-CoV antibodies were negative. Conclusions We provide evidence of effective infection control practices against MERS-CoV transmission in a healthcare facility. Strict infection control precautions can protect HCWs. The optimal infection control measures for MERS-CoV should be further evaluated

Influenza A viral loads in respiratory samples collected from patients infected with pandemic H1N1, seasonal H1N1 and H3N2 viruses

Abstract Background Nasopharyngeal aspirate (NPA), nasal swab (NS), and throat swab (TS) are common specimens used for diagnosis of respiratory virus infections based on the detection of viral genomes, viral antigens and viral isolation. However, there is no documented data regarding the type of specimen that yields the best result of viral detection. In this study, quantitative real time RT-PCR specific for M gene was used to determine influenza A viral loads present in NS, NPA and TS samples collected from patients infected with the 2009 pandemic H1N1, seasonal H1N1 and H3N2 viruses. Various copy numbers of RNA transcripts derived from recombinant plasmids containing complete M gene insert of each virus strain were assayed by RT-PCR. A standard curve for viral RNA quantification was constructed by plotting each Ct value against the log quantity of each standard RNA copy number. Results Copy numbers of M gene were obtained through the extrapolation of Ct values of the test samples against the corresponding standard curve. Among a total of 29 patients with severe influenza enrolled in this study (12 cases of the 2009 pandemic influenza, 5 cases of seasonal H1N1 and 12 cases of seasonal H3N2 virus), NPA was found to contain significantly highest amount of viral loads and followed in order by NS and TS specimen. Viral loads among patients infected with those viruses were comparable regarding type of specimen analyzed. Conclusion Based on M gene copy numbers, we conclude that NPA is the best specimen for detection of influenza A viruses, and followed in order by NS and TS.</p

A Single-Nucleotide Polymorphism in <i>ABCC4</i> Is Associated with Tenofovir-Related Beta2-Microglobulinuria in Thai Patients with HIV-1 Infection

<div><p>Background</p><p>In Thailand, the combined generic anti-retroviral drug stavudine/lamivudine/nevirapine (d4T/3TC/NVP) has been used to treat human immunodeficiency virus (HIV)-infected individuals since 2001. Due to relatively frequent adverse effects, d4T gradually has been replaced with tenofovir disoproxil fumarate (TDF). Although the frequency of adverse drug effects with TDF is lower than that with d4T, TDF is known to induce kidney dysfunction, especially in the proximal tubules. It has been reported that renal tubular transporters, including members of the multi-drug resistant (MDR) protein family, are implicated in tenofovir extrusion and may, therefore, confer susceptibility to TDF-induced kidney tubular dysfunction (KTD). We have explored the association between KTD and polymorphisms in genes that encode adenosine triphosphate-binding cassette (ABC)-type MDRs.</p><p>Methods</p><p>HIV-infected patients receiving TDF-containing antiretroviral regimens for at least one year were enrolled in the study. The levels of beta2-microglobulin in urine and creatinine (Cr) were measured. Three single-nucleotide polymorphisms, <i>ABCC2</i> C-24T (rs717620), <i>ABCC2 G1429A</i> (rs2273697), and <i>ABCC4</i> T4976C (rs1059751), were analyzed using TaqMan SNP genotyping assays.</p><p>Results</p><p>A total of 273 HIV-infected patients were recruited. The median number of years of TDF treatment was 5.04 with interquartile range (IQR) of 3.9–6.7. Despite the length of treatment with TDF, 98.5% patients maintained an estimated glomerular filtration rate (eGFR) of >60 mL/min as calculated by the CKD-EPI formula. Fifty-four patients (19.8%) showed beta2-microglobulinuria (median 2636 μg/g Cr with IQR of 1519–13197 μg/g Cr). The allele frequency of <i>ABCC4</i> T4976C among those 54 patients was 0.602, compared to 0.475 among the 219 remaining patients (p = 0.018).</p><p>Conclusions</p><p>Approximately 20% of HIV-infected patients receiving TDF showed beta2-microglobulinuria. The C allele at position 4976 of the <i>ABCC4</i> gene was associated with beta2-microglobulinuria in this population. This polymorphism may help to identify patients at greater risk for developing TDF-associated KTD.</p></div

Patient-based transcriptome-wide analysis identify interferon and ubiquination pathways as potential predictors of influenza A disease severity.

BACKGROUND: The influenza A virus is an RNA virus that is responsible for seasonal epidemics worldwide with up to five million cases of severe illness and 500,000 deaths annually according to the World Health Organization estimates. The factors associated with severe diseases are not well defined, but more severe disease is more often seen among persons aged >65 years, infants, pregnant women, and individuals of any age with underlying health conditions. METHODOLOGY/PRINCIPAL FINDINGS: Using gene expression microarrays, the transcriptomic profiles of influenza-infected patients with severe (N = 11), moderate (N = 40) and mild (N = 83) symptoms were compared with the febrile patients of unknown etiology (N = 73). We found that influenza-infected patients, regardless of their clinical outcomes, had a stronger induction of antiviral and cytokine responses and a stronger attenuation of NK and T cell responses in comparison with those with unknown etiology. More importantly, we found that both interferon and ubiquitination signaling were strongly attenuated in patients with the most severe outcomes in comparison with those with moderate and mild outcomes, suggesting the protective roles of these pathways in disease pathogenesis. CONCLUSION/SIGNIFICANCES: The attenuation of interferon and ubiquitination pathways may associate with the clinical outcomes of influenza patients

Subjects and time of specimen collection.

<p>Subjects and time of specimen collection.</p

Estimation on the infection rates of the 2009 pandemic influenza in different groups of subjects after the first epide mic wave.

<p>Infection rate is determined by HI titer ≥40 in adults or ≥20 in children.</p