Congenital cytomegalovirus infection is associated with high maternal socio-economic status and corresponding low maternal cytomegalovirus seropositivity.

AIMS: Human cytomegalovirus (CMV) is the leading infectious cause of congenital infection in developed countries. Globally, CMV seropositivity has been associated with low socio-economic status (SES); however, Australian data are lacking. Therefore, we examined the association between SES and CMV seroprevalence in children and pregnant women. METHODS: Three groups were examined: 1, a prospective cohort of Australian children aged 0-15 years (n = 220); 2, a clinic-based sample of pregnant women (n = 778); and 3, a case series of infants and children (n = 219) with symptomatic congenital CMV infection. SES was determined using a postcode-based score from the Australian Bureau of Statistics.Group 1 was recruited from endocrinology clinics and follow-up at Prince of Wales Hospital and Children's Hospital at Westmead. Group 2 was recruited at the Royal Hospital for Women. Congenitally infected infants were identified through the Australian Paediatric Surveillance Unit. RESULTS: CMV seroprevalence among all children was 20% (95% confidence interval (CI) 15-25%), and there was no association with SES (P = 0.58). Seroprevalence among pregnant women was 57% (53-60%), and higher rates of CMV seropositivity were associated with lower SES (P < 0.001). More congenital CMV cases were reported in the highest socio-economic groups (55%) than the lowest (9%) (P < 0.001). CONCLUSIONS: A marked socio-economic gradient in CMV seroprevalence is evident in Australian pregnant women and cases of congenital CMV but not in unselected Australian children. These findings highlight the importance of a community-wide approach to CMV awareness and the potential for hygienic measures to reduce the burden of congenital CMV in Australia.

Detecting human papillomavirus in ocular surface diseases.

PURPOSE: Human papillomavirus (HPV) infection has been implicated as a possible inducing factor for benign and neoplastic ocular surface diseases such as pterygia and ocular-surface squamous neoplasia (OSSN). However, the wide range in HPV prevalence previously reported for both diseases adds controversy to, and highlights the limitations of, this field. The aim of this study was to determine the prevalence of HPV in pterygia and OSSN and to devise a standardized approach for detecting viral DNA in ocular tissue samples. METHODS: DNA was extracted from a variety of specimens (n = 160), including formalin-fixed paraffin-embedded tissue shavings, fresh tissue, and cultured cells. Nested PCR for HPV with consensus and subtype-specific primers was used to detect viral DNA. Confirmatory assays, including molecular sequencing, histology, and immunohistochemistry for HPV E6 protein and p16 were also performed. RESULTS: HPV was not detected in pterygia or normal conjunctiva. However, 6.5% (3/46) of OSSN samples were HPV-positive by PCR, sequencing, and immunohistochemistry. Positive cases were all squamous cell carcinoma of the conjunctiva (SCCC), the most severe form of OSSN, representing 12.5% (3/24) of SCCCs in our cohort. HPV-16 was the genotype identified in each case and this correlated with the presence of koilocytes and intense immunoreactivity for p16. Our study found no association between pterygia and OSSN with other oncogenic viruses, such as EBV or CMV, as they were just as prevalent in normal conjunctiva. CONCLUSIONS: The low prevalence of HPV-16 in ocular surface disease suggests infection is not a cause but a cofactor in disease development.

A pilot study to determine the feasibility of collecting amniotic fluid samples from women during labour and measuring amniotic fluid lactate at point of care.

BACKGROUND: The level of lactate in amniotic fluid may provide useful clinical information when assessing progress of a woman's labour and if so, a rapid, reliable method to assess amniotic fluid lactate is required in order to be clinically relevant. However, measuring lactate levels in amniotic fluid, using portable, handheld lactate meters may be less accurate than reference laboratory instruments designed to measure lactate levels in aqueous solutions. Prior to conducting a large study, we assessed recruitment, consent and sampling procedures, and the accuracy of a handheld lactate meter to measure lactate in amniotic fluid. We compared amniotic fluid lactate results obtained using the hand held Lactate Pro (Arkray) to results obtained using reference laboratory methods ABX Pentra 400 (Horiba). RESULTS: We recruited 35 nulliparous women during their antenatal hospital visits and tested amniotic fluid samples collected from 20 labouring women. The handheld Lactate Pro meter was found accurate from 9-20 mmol/L with a Passing & Bablok regression of y = 0.18 + 0.97x (95% CI 0.76-1.45). Amniotic fluid lactate results remained reliable in the presence of potential contaminants commonly encountered during labour; obstetric lubricant, blood and meconium. CONCLUSION: The measurement of amniotic fluid lactate using the Lactate Pro meter was reliable compared to reference laboratory methods for measuring lactate levels in amniotic fluid. The pilot study enabled the refinement of information, recruitment, consenting and sampling procedures prior to commencing a large cohort study.

Human cytomegalovirus-induces cytokine changes in the placenta with implications for adverse pregnancy outcomes.

Human cytomegalovirus (CMV) infection of the developing fetus can result in adverse pregnancy outcomes including death in utero. Fetal injury results from direct viral cytopathic damage to the CMV-infected fetus, although evidence suggests CMV placental infection may indirectly cause injury to the fetus, possibly via immune dysregulation with placental dysfunction. This study investigated the effects of CMV infection on expression of the chemokine MCP-1 (CCL2) and cytokine TNF-α in placentae from naturally infected stillborn babies, and compared these changes with those found in placental villous explant histocultures acutely infected with CMV ex vivo. Tissue cytokine protein levels were assessed using quantitative immunohistochemistry. CMV-infected placentae from stillborn babies had significantly elevated MCP-1 and TNF-α levels compared with uninfected placentae (p = 0.001 and p = 0.007), which was not observed in placentae infected with other microorganisms (p = 0.62 and p = 0.71) (n = 7 per group). Modelling acute clinical infection using ex vivo placental explant histocultures showed infection with CMV laboratory strain AD169 (0.2 pfu/ml) caused significantly elevated expression of MCP-1 and TNF-α compared with uninfected explants (p = 0.0003 and p<0.0001) (n = 25 per group). Explant infection with wild-type Merlin at a tenfold lower multiplicity of infection (0.02 pfu/ml), caused a significant positive correlation between increased explant infection and upregulation of MCP-1 and TNF-α expression (p = 0.0001 and p = 0.017). Cytokine dysregulation has been associated with adverse outcomes of pregnancy, and can negatively affect placental development and function. These novel findings demonstrate CMV infection modulates the placental immune environment in vivo and in a multicellular ex vivo model, suggesting CMV-induced cytokine modulation as a potential initiator and/or exacerbator of placental and fetal injury.

Human cytomegalovirus infection is detected frequently in stillbirths and is associated with fetal thrombotic vasculopathy.

BACKGROUND: Human cytomegalovirus (CMV) is the most common congenital infection in developed countries and is a known cause of intrauterine fetal death. We examined CMV infection in stillbirths and the relationship with histopathological findings at autopsy. METHODS: We collected liver, kidney, and placenta specimens from 130 stillbirths. CMV DNA and protein were detected using polymerase chain reaction and immunohistochemistry, along with routine autopsy of stillborn infants. RESULTS: Overall, CMV DNA was detected in 15% of singleton, >20-week stillborn infants. CMV DNA was detected in kidney (9%), liver (11%), and placenta (5%) specimens, with 75% of infections confirmed by immunohistochemistry. Fetal thrombotic vasculopathy was the only histopathological abnormality associated with CMV infection (in 60% CMV-infected vs 28% uninfected stillbirths P = .010). CONCLUSIONS: Stillbirth has multiple etiologies. However, the detection of CMV DNA in 15% of fetal tissues or placentae suggests a strong association between CMV infection in pregnancy and stillbirth. Molecular testing during postmortem investigation has an important role to determine the contribution of CMV infection.

Outbreaks of pandemic (H1N1) 2009 and seasonal influenza A (H3N2) on cruise ship.

To determine the extent and pattern of influenza transmission and effectiveness of containment measures, we investigated dual outbreaks of pandemic (H1N1) 2009 and influenza A (H3N2) that had occurred on a cruise ship in May 2009. Of 1,970 passengers and 734 crew members, 82 (3.0%) were infected with pandemic (H1N1) 2009 virus, 98 (3.6%) with influenza A (H3N2) virus, and 2 (0.1%) with both. Among 45 children who visited the ship's childcare center, infection rate for pandemic (H1N1) 2009 was higher than that for influenza A (H3N2) viruses. Disembarked passengers reported a high level of compliance with isolation and quarantine recommendations. We found 4 subsequent cases epidemiologically linked to passengers but no evidence of sustained transmission to the community or passengers on the next cruise. Among this population of generally healthy passengers, children seemed more susceptible to pandemic (H1N1) 2009 than to influenza (H3N2) viruses. Intensive disease control measures successfully contained these outbreaks.

Enhanced diagnosis of pandemic (H1N1) 2009 influenza infection using molecular and serological testing in intensive care unit patients with suspected influenza.

During the 2009 outbreak of pandemic (H1N1) 2009 influenza (pH1N1) in Australia, acute and convalescent serum specimens were collected from 33 patients with severe respiratory disease admitted to intensive care units. Using hemagglutination inhibition of pH1N1, 29 paired serum samples showed significant increases in specific antibody titers. Of these 29 patients, 18 had pH1N1 RNA detected by routine nucleic acid testing. These results indicate that up to one-third of pH1N1 cases may not have laboratory confirmation of infection unless serological testing is included for suspected cases.

Successful valganciclovir treatment of post-transplant cytomegalovirus infection in the presence of UL97 mutation N597D.

Mutations in the human cytomegalovirus (CMV) UL97 protein kinase are the most common mechanism of ganciclovir (GCV) resistance in the clinical setting. A CMV strain with a previously unrecognized UL97 mutation N597D was identified in the blood of a heart transplant recipient who experienced a persistent CMV infection with high viral loads accompanying pain and fever while receiving valganciclovir (valGCV) therapy. The N597D mutation was transferred by mutagenesis to an antiviral sensitive CMV strain for analysis of antiviral susceptibility by standardized phenotypic assay. Recombinant phenotyping showed N597D conferred a less than twofold increase in GCV IC(50) compared to the sensitive control strain. Despite the presence of this mutation, valGCV eventually resolved the infection after 6 weeks of therapy. A subsequent CMV reactivation was also responsive to valganciclovir. This case illustrates the diversity of UL97 mutations in the codon segment 590-607 usually associated with GCV resistance, with some mutations producing minimal levels of resistance that do not preclude a therapeutic response to the drug. Accurate interpretation of genotypic test results ultimately requires experimental determination of the level of resistance conferred by newly discovered UL97 mutations.

Emergence and persistence of multiple antiviral-resistant CMV strains in a highly immunocompromised child.

BACKGROUND: The emergence of human cytomegalovirus (CMV) antiviral resistance plays a significant role in disease progression in immunocompromised patients who have received antiviral therapy. OBJECTIVES: To determine the pattern of antiviral-resistant CMV strains in a highly immunocompromised child. STUDY DESIGN: Retrospective specimens of blood and urine were analysed using PCR-sequencing to identify antiviral-resistant CMV strains containing UL97 or UL54 mutations. RESULTS: CMV strains resistant to antiviral agents contributed to disease in a bone marrow transplant recipient with X-linked severe combined immunodeficiency (SCID) treated with ganciclovir (GCV) and foscarnet (FOS). Retrospective analyses detected GCV-resistant CMV (L595S) in a specimen taken after disease progression. This GCV-resistant CMV strain persisted for 1 year, after which time it was no longer detected even though the patient continued to receive GCV. A FOS-resistant strain (T700A) then emerged even though no FOS had been administered in the preceding year. CONCLUSION: The detection of antiviral-resistant CMV did not follow the patterns found in other patients tested for antiviral resistance, including emergence of a FOS-resistant strain in the absence of antiviral-selective pressure. These findings indicate the patient's underlying immunosuppressive condition should be considered for diagnosis and management of resistant CMV.