Detection of IgG3 antibodies specific to the human immunodeficiency virus type 1 (HIV-1) p24 protein as marker for recently acquired infection.

Reducing the risk of human immunodeficiency virus type 1 (HIV-1) transmission is still a public health priority. The development of effective control strategies relies on the quantification of the effects of prophylactic and therapeutic measures in disease incidence. Although several assays can be used to estimate HIV incidence, these estimates are limited by the poor performance of these assays in distinguishing recent from long-standing infections. To address such limitation, we have developed an assay to titrate p24-specific IgG3 antibodies as a marker of recent infection. The assay is based on a recombinant p24 protein capable to detect total IgG antibodies in sera using a liquid micro array and enzyme-linked immunosorbent assay. Subsequently, the assay was optimised to detect and titrate anti-p24 IgG3 responses in a panel of sequential specimens from seroconverters over 24 months. The kinetics of p24-specific IgG3 titres revealed a transient peak in the 4 to 5-month period after seroconversion. It was followed by a sharp decline, allowing infections with less than 6 months to be distinguished from older ones. The developed assay exhibited a mean duration of recent infection of 144 days and a false-recent rate of ca. 14%. Our findings show that HIV-1 p24-specific IgG3 titres can be used as a tool to evaluate HIV incidence in serosurveys and to monitor the efficacy of vaccines and other transmission control strategies.

Establishment and cryptic transmission of Zika virus in Brazil and the Americas.

Transmission of Zika virus (ZIKV) in the Americas was first confirmed in May 2015 in northeast Brazil. Brazil has had the highest number of reported ZIKV cases worldwide (more than 200,000 by 24 December 2016) and the most cases associated with microcephaly and other birth defects (2,366 confirmed by 31 December 2016). Since the initial detection of ZIKV in Brazil, more than 45 countries in the Americas have reported local ZIKV transmission, with 24 of these reporting severe ZIKV-associated disease. However, the origin and epidemic history of ZIKV in Brazil and the Americas remain poorly understood, despite the value of this information for interpreting observed trends in reported microcephaly. Here we address this issue by generating 54 complete or partial ZIKV genomes, mostly from Brazil, and reporting data generated by a mobile genomics laboratory that travelled across northeast Brazil in 2016. One sequence represents the earliest confirmed ZIKV infection in Brazil. Analyses of viral genomes with ecological and epidemiological data yield an estimate that ZIKV was present in northeast Brazil by February 2014 and is likely to have disseminated from there, nationally and internationally, before the first detection of ZIKV in the Americas. Estimated dates for the international spread of ZIKV from Brazil indicate the duration of pre-detection cryptic transmission in recipient regions. The role of northeast Brazil in the establishment of ZIKV in the Americas is further supported by geographic analysis of ZIKV transmission potential and by estimates of the basic reproduction number of the virus.

Development of potent class II transactivator gene delivery systems capable of inducing de novo MHC II expression in human cells, in vitro and ex vivo.

Class II transactivator (CIITA) induces transcription of major histocompatibility complex (MHC) II genes and can potentially be used to improve genetic immunotherapies by converting non-immune cells into cells capable of presenting antigens to CD4+ T cells. However, CIITA expression is tightly controlled and it remains unclear whether distinct non-immune cells differ in this transactivator regulation. Here we describe the development of gene delivery systems capable of promoting the efficient CIITA expression in non-immune cell lines and in primary human cells of an ex vivo skin explant model. Different human cell types undergoing CIITA overexpression presented high-level de novo expression of MHC II, validating the delivery systems as suitable tools for the CIITA evaluation as a molecular adjuvant for gene therapies.

Development of a DNA-based vaccine strategy against bovine papillomavirus infection, involving the E5 or L2 gene.

Papillomaviruses are known to cause tumor lesions, generally benign, in epithelial tissues of diverse organisms; these lesions may progress to cancer under suitable conditions. Bovine papillomavirus (BPV) can cause urinary bladder cancer and cancer of the upper gastrointestinal tract. Furthermore, BPV1 and BPV2 are implicated in the development of tumors in equids. Many studies with animal models clearly demonstrate that DNA vaccines are very effective tools in controlling viral infections, providing strong humoral and cellular immune responses. In this study, we have described the development of two vaccine constructs for the control of diseases caused by BPV. The 1st strategy is prophylactic and is based on the L2 gene; the 2nd is therapeutic and is based on the E5 gene. Vaccine constructs were obtained and evaluated in vitro in mammalian cells. The results show the occurrence of E5 and L2 transcription and viral protein production. These results confirm the functionality of the vaccine constructs in mammalian cells. This is the 1st step in the development of a DNA-based vaccine strategy for the control and/or treatment of diseases caused by BPV.

Diagnosis of Wuchereria bancrofti infection by the polymerase chain reaction using urine and day blood samples from amicrofilaraemic patients.

A sensitive and specific polymerase chain reaction (PCR) based on a highly repeated deoxyribonucleic acid (DNA) sequence (188 bp; SspI repeat) was tested for the detection of Wuchereria bancrofti DNA in blood and urine samples collected during the day from individuals in Coque, Recife, Brazil, an endemic area for W. bancrofti. All microfilaraemic individuals were also positive by PCR, irrespective of the samples used. The PCR system was capable of detecting W. bancrofti DNA in amicrofilaraemic individuals: c. 93% were positive by PCR when day blood samples were used and 59.7% when urine samples collected at 07:00 were used. Thus, nocturnally periodic W. bancrofti infection can be detected in blood samples collected during the day, which is convenient for large-scale screening. In addition, non-invasive urine collection provided suitable samples for PCR, which is clearly advantageous for preliminary mass diagnosis.