ABSTRACT
The aim of this study was to determine whether a patient's endocervical swab specimen can be transported in first void urine (FVU) as combined specimens for the detection of Mycoplasma genitalium by real-time PCR. The study also compared two different DNA extraction methods for observation of possible PCR inhibition. Three specimens, one endocervical swab specimen transported in 2-SP medium, one endocervical swab specimen transported in FVU and a FVU specimen, were collected from 329 women. All sample types underwent manual DNA extraction whereas in the DNA extraction study, 329 endocervical swab specimens transported in FVU were subjected to both manual Chelex and automated BioRobot M48 DNA extraction. A total of 100 endocervical swab specimens transported in FVU from patients PCR-negative for M. genitalium in the study were used in the PCR inhibition analysis. M. genitalium was detected in 25/329 (7.6 %) women. The endocervical swab specimens transported in 2-SP medium and transported in FVU were positive for M. genitalium in 17/25 (68 %) and 24/25 (96 %) women, respectively. The FVU specimens alone were positive for M. genitalium in 22/25 (88 %) women. In the DNA extraction study, M. genitalium DNA was detected in 24/329 (7.3 %) and 28/329 (8.5 %) of endocervical swab specimens transported in FVU subjected to manual Chelex extraction and automated BioRobot M48 extraction, respectively. Partial PCR inhibition was detected in 6 % of samples subjected to manual Chelex extraction whereas no inhibition was detected with the automated BioRobot M48 extraction. Thus endocervical swab specimens transported in FVU demonstrate higher sensitivity than FVU specimens only and have considerably increased sensitivity compared with endocervical swab specimens transported in 2-SP medium for detection of M. genitalium DNA. Moreover, automated BioRobot M48 extraction was shown to be superior to a crude manual Chelex extraction, leaving no PCR inhibition and giving a slightly higher DNA yield and/or better sensitivity.
Subject(s)
Cervix Uteri/microbiology , Mycoplasma Infections/diagnosis , Mycoplasma genitalium/isolation & purification , Polymerase Chain Reaction , Specimen Handling/methods , Adolescent , Adult , Aged , DNA, Bacterial/isolation & purification , Female , Humans , Middle Aged , Mycoplasma Infections/urine , Vaginal Diseases/diagnosis , Vaginal Diseases/microbiologyABSTRACT
In this study, we tested the hypothesis that exposure to a maternal infection during fetal life can lead to the appearance of alterations in the brain later in life. C57BL/6 mice were infected intranasally with influenza A/WSN/33 virus on day 14 of gestation. The levels of transcripts encoding neuroleukin and fibroblast growth factor 5 were significantly elevated in the brains of the virus-exposed offspring at 90 and 280 days of age, but not at earlier time-points. For neuroleukin, this difference could also be observed at the protein level. Thus, a maternal influenza A virus infection can give rise to alterations in gene expression in the brain that become apparent only after a prepubertal latency period.
Subject(s)
Brain/metabolism , Central Nervous System Viral Diseases/metabolism , Gene Expression Regulation, Developmental/physiology , Maternal Exposure , Prenatal Exposure Delayed Effects , Animals , Brain/virology , Central Nervous System Viral Diseases/virology , Female , Humans , Influenza A virus/genetics , Influenza A virus/metabolism , Influenza, Human/genetics , Influenza, Human/metabolism , Mice , Mice, Inbred C57BL , PregnancyABSTRACT
Invasion and persistence of the neuroadapted influenza virus A/WSN/33 in the mouse olfactory system was studied. WSN/33 instilled intranasally infected neurons in the olfactory epithelium and was transported in axons to the olfactory bulbs in wild type mice that survived the infection. In adult mice lacking the recombination activating gene 1 (RAG-1-/-), infected neurons occurred in the olfactory bulbs for 22-65 days after which the mice developed a rapidly progressive lethal infection affecting neurons in olfactory projection pathways, i.e. primary olfactory cortex, raphe in upper brainstem and hypothalamus. Adult mice without genes for interferon (IFN)-alpha/beta receptor, IFN-gamma receptor, inducible nitric oxide synthase (iNOS), IgH, the transporter associated with antigen processing 1 (TAP1), and natural killer cell-depleted mice, all survived the infection. Viral RNA was found in the olfactory bulbs in more than 80 per cent of the surviving iNOS-/-, IFN-gamma receptor-/-, and TAP1-/- mice. Taken together, this study shows that influenza A virus can invade the brain through the olfactory pathways and that the cellular immune responses prevent establishment of persistent infections in the olfactory bulbs. Furthermore, innate responses in olfactory bulbs may for a period of time keep the infection under control.
Subject(s)
Influenza A virus/pathogenicity , Olfactory Bulb/virology , Orthomyxoviridae Infections/virology , ATP Binding Cassette Transporter, Subfamily B, Member 2 , ATP-Binding Cassette Transporters/genetics , ATP-Binding Cassette Transporters/immunology , Adaptation, Physiological , Animals , Base Sequence , DNA, Complementary/genetics , DNA, Viral/genetics , Genes, RAG-1 , Immunity, Cellular , Influenza A virus/genetics , Influenza A virus/physiology , Membrane Proteins , Mice , Mice, Inbred C57BL , Mice, Knockout , Nitric Oxide Synthase/deficiency , Nitric Oxide Synthase/genetics , Nitric Oxide Synthase Type II , Olfactory Bulb/immunology , Orthomyxoviridae Infections/genetics , Orthomyxoviridae Infections/immunology , Receptor, Interferon alpha-beta , Receptors, Interferon/deficiency , Receptors, Interferon/genetics , Interferon gamma ReceptorABSTRACT
Epidemiological studies have indicated an association between influenza A virus infections during fetal life and neuropsychiatric diseases. To study the potential for influenza A virus infections to cause nervous system dysfunctions, we describe a mouse model using intranasal instillation of the mouse neuroadapted influenza A/WSN/33 strain in pregnant mice. Viral RNA and nucleoprotein were detected in fetal brains and the viral RNA persisted for at least 90 days of postnatal life. We have, thus, obtained evidence for transplacental passage of influenza virus in mice and the persistence of viral components in the brains of these animals into young adulthood.
