Effects of remdesivir on SARS-CoV-2 viral dynamics and mortality in viraemic patients hospitalized for COVID-19.

BACKGROUND: Studies on the antiviral effects of remdesivir have shown conflicting results. SARS-CoV-2 viraemia could identify patients in whom antiviral treatment may be particularly beneficial. OBJECTIVES: To investigate antiviral effects and clinical outcomes of remdesivir treatment in viraemic patients. METHODS: Viraemic patients hospitalized for COVID-19 with ratio of arterial oxygen partial pressure to fractional inspired oxygen of ≤300, symptom duration ≤10 days, and estimated glomerular filtration rate ≥30 mL/min were included in a cohort. The rate of serum viral clearance and serum viral load decline, 60 day mortality and in-hospital outcomes were estimated. A subgroup analysis including patients with symptom duration ≤7 days was performed. RESULTS: A total of 318 viraemic patients were included. Thirty-three percent (105/318) received remdesivir. The rate of serum viral clearance [subhazard risk ratio (SHR) 1.4 (95% CI 0.9-2.0), P = 0.11] and serum viral load decline (P = 0.11) were not significantly different between remdesivir-treated patients and controls. However, the rate of serum viral clearance was non-significantly higher [SHR 1.6 (95% CI 1.0-2.7), P = 0.051] and the viral load decline was faster (P = 0.03) in remdesivir-treated patients with symptom duration ≤7 days at admission. The 60 day mortality [HR 1.0 (95% CI 0.6-1.8), P = 0.97] and adverse in-hospital outcomes [OR 1.4 (95% CI 0.8-2.4), P = 0.31] were not significantly different between remdesivir-treated patients and controls. CONCLUSIONS: Remdesivir treatment did not significantly change the duration of SARS-CoV-2 viraemia, decline of serum viral load, 60 day mortality or in-hospital adverse outcomes in patients with ≤10 days of symptoms at admission. Remdesivir appeared to reduce the duration of viraemia in a subgroup of patients with ≤7 days of symptoms at admission.

Correlation of SARS-CoV-2 Nasopharyngeal CT Values With Viremia and Mortality in Adults Hospitalized With COVID-19.

Background: Both severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) viremia and nasopharyngeal viral load have been suggested to be predictors of unfavorable outcome in coronavirus disease 2019 (COVID-19). This study aimed to investigate whether nasopharyngeal viral load is correlated with viremia and unfavorable outcome. Methods: The presence of SARS-CoV-2 RNA was determined in paired nasopharyngeal and serum samples collected at admission from patients hospitalized for COVID-19. Standardized cycle threshold values (CT values) were used as an indicator of viral load. An adjusted logistic regression was used to estimate the risk of viremia at different nasopharyngeal CT values. A Cox regression was used to estimate the risk of 60-day mortality. Results: A total of 688 patients were included. Viremia at admission was detected in 63% (146/230), 46% (105/226), and 31% (73/232) of patients with low, intermediate, and high nasopharyngeal CT values. The adjusted odds ratios of being viremic were 4.4 (95% CI, 2.9-6.8) and 2.0 (95% CI, 1.4-3.0) for patients with low and intermediate CT values, compared with high CT values. The 60-day mortality rate was 37% (84/230), 15% (36/226), and 10% (23/232) for patients with low, intermediate, and high nasopharyngeal CT values at admission, respectively. Adjusted hazard ratios were 2.6 (95% CI, 1.6-4.2) and 1.4 (95% CI, 0.8-2.4) for patients with low and intermediate CT values compared with high CT values. Conclusions: There was a dose-dependent correlation between nasopharyngeal CT values and viremia at admission for COVID-19. Moreover, there was an increased risk of 60-day mortality for patients with low, compared with high, nasopharyngeal CT values.

Cytomegalovirus infection and neonatal outcome in extremely preterm infants after freezing of maternal milk.

BACKGROUND: Cytomegalovirus (CMV) infection acquired from breast milk can cause serious illness in extremely preterm (EPT) infants (<28 weeks). Some neonatal centers freeze maternal milk (MM) to prevent CMV transmission; however, this practice is controversial. In this study, we assessed the CMV transmission rate and neonatal outcome in EPT infants after routine freezing of all MM. METHODS: EPT infants (n = 140) and their mothers were randomized to the intervention group (only freeze-thawed MM) or the control group (combined fresh and freeze-thawed MM). Freeze-thawed MM was frozen at -20°C for ≥3 days before thawing. Mothers had serological tests for CMV, and MM was analyzed for CMV by polymerase chain reaction and CMV culture. Infants underwent CMV screening with urine analysis by CMV-polymerase chain reaction and CMV culture until 12 weeks of age. RESULTS: Congenital CMV infection was detected in 2% of screened infants. The CMV transmission rate in infants fed with CMV-DNA positive milk was 8% (3 of 37) in the intervention group and 6% (2 of 33) in controls. All infants infected by CMV were asymptomatic. The final per-protocol analysis included 56 infants in the intervention group and 65 controls. Neonatal mortality was comparable between the groups (7% vs. 6%). Neonatal morbidity was similar, except for late onset Candida sepsis, which was more frequent in the controls (12% vs. 0%). CONCLUSIONS: Routine freezing of all MM did not affect the rate of CMV transmission but may help to prevent fungal sepsis in EPT infants. This observation merits further investigation.

Children with multiple viral respiratory infections are older than those with single viruses.

AIM: To study the clinical impact of multiple viral respiratory infections compared to single infections. METHODS: Demographic data from 37 multiple infection periods in children <5 years of age were compared to data from 193 episodes with single infections. Clinical data derived from patient records of the multiple infection episodes were further compared to data from 93 matched control episodes with single infections. RESULTS: The mean age of patients with multiple viral findings was 12.7 months, compared to 5.7 months for those with single findings (p < 0.01). Wheezing was the most common diagnosis in both groups, except among children who were only infected with the coronavirus. No differences were found regarding duration of hospitalisation, oxygen treatment or admittance to the intensive care unit. CONCLUSION: Children with multiple viral findings in their respiratory secretions were older than those with a single detected virus. Otherwise, no major differences in comorbidity, presentation or clinical outcome were observed between the two groups.

Use of oral fluid to examine the molecular epidemiology of varicella zoster virus in the United Kingdom and continental Europe.

We investigated oral fluid (OF) as an alternative to sampling of rashes for varicella zoster virus (VZV) genotyping and further characterized VZV clade prevalence in the United Kingdom and Europe. VZV was detected in up to 91% of OF specimens. Paired OF and vesicle fluid samples contained identical VZV clades. While clades 1 and 3 were the most prevalent across the United Kingdom and Europe, in Western Europe, clade 5 viruses were circulating. Viruses from the same outbreak belonged to different clades, but no clade was associated with a severe-disease phenotype. OF is suitable and convenient for large-scale molecular epidemiological studies of VZV.

Genotyping of human papillomavirus in triaging of low-grade cervical cytology.

OBJECTIVE: The objective of the study was to evaluate whether typing of human papillomavirus (HPV) among women with low-grade cervical cytology can improve the ability to identify women with cervical cancer or cervical intraepithelial neoplasia grade III (CIN III or worse). STUDY DESIGN: A total of 1595 women with low-grade cervical cytology participating in a randomized implementation trial of HPV triaging using Hybrid Capture II were also HPV genotyped and CIN III or worse predictive values evaluated. RESULTS: HPV 16 was detected in 57% of cases with CIN III or worse but only among 24% of all tested women. Testing for the 3 HPV types with highest risk (HPV16/31/33) detected 77% of CIN III or worse, with 36% of women testing positive. Positivity for the other high-risk HPV types had a decreased risk for CIN III or worse. CONCLUSION: Different high-risk HPV types confer different risks for the presence of CIN III or worse, implying that HPV genotyping could be useful for the optimization of triaging strategies.

Randomized healthservices study of human papillomavirus-based management of low-grade cytological abnormalities.

Human papillomavirus (HPV)-based management of women with borderline atypical squamous cells of undetermined significance (ASCUS) or mildly abnormal cervical intraepithelial neoplasia (CINI) cervical cytology has been extensively studied in the research setting. We wished to assess safety and health care resource use of a real-life health care policy using HPV triaging. All 15 outpatient clinics involved in the organized population-based screening program in Stockholm, Sweden screening program were randomized to either continue with prior policy (colposcopy of all women with ASCUS/CINI) or to implement a policy with HPV triaging and colposcopy only of HPV-positive women. The trial enrolled the 3,319 women who were diagnosed with ASCUS (n = 1,335) or CINI (n = 1,984) in Stockholm during 17th March 2003 to 16th January 2006. Detection of high-grade cervical lesions (CINII+) and health care cost consumption was studied by registry linkages. The proportion of histopathology-verified CINII+ was similar for the two policies (395 of 1,752 women (22.5%; 95% Confidence interval [CI]: 20.6-24.6%) had CINII+ diagnosed with HPV triaging policy, 318 of 1,567 women (20.3%; 95%CI: 18.3-22.4%) had CINII+ with colposcopy policy). Sixty-four percent of women with ASCUS and 77% of women with CINI were HPV positive. HPV-positivity was age-dependent, with 81% of women below 35 years of age and 44% of women above 45 years of age testing HPV-positive. HPV triaging was cost-effective only above 35 years of age. In conclusion, a real-life randomized healthservices study of HPV triaging of women with ASCUS/CINI demonstrated similar detection of CINII+ as colposcopy of all women.

Development and implementation of a molecular diagnostic platform for daily rapid detection of 15 respiratory viruses.

Acute respiratory tract infections are caused by a large number of viruses. Diagnostic methods have until recently been available only for a limited number of these viruses. With the objective to achieve sensitive assays for all respiratory viruses, a rational workflow in the laboratory, and a short turn-around time, a real-time PCR diagnostic platform for daily rapid detection of 15 respiratory viruses was developed. The system was evaluated on 585 stored nasopharyngeal aspirates from hospitalized children. Previous analysis by immunofluorescence and virus isolation identified viruses in 37% of the samples while the new PCR diagnostic panel detected 57% virus positive samples. The new platform was introduced in the laboratory in October 2007 and has then fully replaced the standard immunofluorescence assay for rapid detection of viruses and virus isolation.

Infection with Parvovirus B19 and Herpes viruses in early pregnancy and risk of second trimester miscarriage or very preterm birth.

We investigated whether infections with Parvovirus B19 and Herpes viruses in early pregnancy increase risks of second trimester miscarriage or delivery before 32 gestational weeks. Blood samples taken in early pregnancy were analyzed for Parvovirus B19 or Herpes viruses. Viremia was found in blood samples of 11 (4.7%) women with second trimester miscarriage and 10 (3.7%) women with very preterm birth, compared to 5 (1.7%) women who delivered at term, corresponding to adjusted odds ratios [95% CI] of 3.32 [0.93, 11.8] and 2.21 [0.71, 6.84], respectively. In stratified analyses, Parvovirus B19 viremia was associated with adjusted odds ratios of 3.76 [0.77, 18.3] for second trimester miscarriage and 2.66 [0.64, 11.1] for very preterm birth. Corresponding odds ratios for Human Herpes virus 6 viremia was 2.52 [0.33, 19.5] and 1.08 [0.14, 8.08], respectively. In conclusion, this study lends some support to the hypothesis that women with viremia in early pregnancy may face an increased risk of second trimester miscarriage or very preterm birth. Studies with larger sample sizes are needed.

High diagnostic yield by CSF-PCR for entero- and herpes simplex viruses and TBEV serology in adults with acute aseptic meningitis in Stockholm.

Acute aseptic meningitis (AAM) affects 10-20/100,000 inhabitants per years in Sweden. Up to the beginning of the 1980s the diagnoses were made by virus isolation and/or determination of viral antibodies in serum. The development of PCR for detection of viruses in CSF samples has increased the sensitivity and diagnostic efficiency considerably. We investigated the aetiology of AAM and the diagnostic efficiency in an adult population in Stockholm, using a limited first-line combination of microbiological assays. CSF and serum samples, consecutively collected in 419 patients with clinical symptoms of AAM in northern Stockholm during 1999-2004, were included. PCR assays for herpes simplex virus (HSV) DNA and enterovirus (EV) RNA in the CSF as well as ELISA for IgM in serum to tick-borne encephalitis virus (TBEV) were performed routinely. A viral diagnosis was obtained in 255 of the 419 cases (62%) with these routinely performed assays. Clinical findings in combination with additional diagnostic tests resulted in an overall aetiological yield of 72%. EV was the major causative agent (27%) followed by TBEV (21%) and HSV-2 (19%). We conclude that consistent use of CSF-PCR for EV and HSV and TBEV serology established a diagnosis in the majority of AAM patients.

Molecular epidemiology and genetic variability of respiratory syncytial virus (RSV) in Stockholm, 2002-2003.

The epidemiology and genetic variability of circulating respiratory syncytial virus (RSV) strains in Stockholm during the season 2002-2003 were studied in consecutive RSV isolates derived from respiratory samples and diagnosed in the laboratory. Two hundred thirty-four viruses were sequenced. The samples were mainly from children under 1 year old (79%). The phylogeny of the N-terminal part of the G gene was studied after amplification and sequencing. One hundred fifty-two viruses belonged to subgroup B and 82 to subgroup A. The subgroup A viruses could be further divided into genotypes GA2 (25) and GA5 (57) and the subgroup B viruses into GB3 (137) and SAB1 (15) strains. These strains clustered with subgroup A and subgroup B strains from Kenya from the same period, as well as with strains from Great Britain from 1995 to 1998. The dominance of subgroup B strains in Stockholm during 2002-2003 is in agreement with findings from other parts of the world during the same years. Only two genotypes of subgroup A, GA2 and GA5, were circulating during this time, and GA2 has been circulating in Sweden for more than 20 years. Consecutive strains from the same individual displayed no variability in the sequenced region, which was also true of strains that had been passaged in cell cultures.

High occurrence of a new variant of Chlamydia trachomatis escaping diagnostic tests among STI clinic patients in Stockholm, Sweden.

BACKGROUND: In 2006, a genetic variant of Chlamydia trachomatis not detectable with the most commonly used diagnostic tests was identified. Initial reports suggested that as many as 10% to 13% of all chlamydia cases would have remained undiagnosed. The aim of the study was to find the occurrence and clinical findings of this genetic variant among a high-risk population in Stockholm, Sweden. METHODS: Samples were analyzed using the Cobas TaqMan CT test (Roche Diagnostics). To detect the new variant, an additional PCR-analysis, artus C. trachomatis LC MOMP PCR Kit (Qiagen) was performed on all negative samples. Positive results in the artus test were confirmed by a mutant specific PCR. Clinical data were retrospectively collected from medical records. RESULTS: Among 1009 samples analyzed, 115 were positive for C. trachomatis and among those, 27 were found to belong to the genetic altered strain. This variant constituted 23% of all chlamydia cases diagnosed, and 29% were found in the age group 20 to 29 years. Women with the new variant were younger and had more often performed another chlamydia test within the previous 6 months compared with those infected with the wild type. CONCLUSION: These results indicate that a large number of sexually active individuals might be infected despite a negative chlamydia test, thus facilitating a rapid transmission of the new variant. Accordingly, it is of great importance to be aware of limitations of the diagnostic methods used.

Increased detection rate in diagnosis of herpes simplex virus type 2 meningitis by real-time PCR using cerebrospinal fluid samples.

Efficient and sensitive diagnostic methods are needed in the management of virus infections in the central nervous system. There is a demand for an evaluation of the sensitivity of PCR methods for early diagnosis of meningitis due to herpes simplex type 2 (HSV-2) and varicella-zoster virus (VZV). The objective of this study was to evaluate real-time PCR in the detection of HSV-2 and VZV DNA from cerebrospinal fluid (CSF) for etiological diagnoses in clinically well-characterized cases of primary and recurrent aseptic meningitis. Samples from 110 patients, 65 of whom were diagnosed with or were strongly suspected of having HSV-2 meningitis and 45 with acute aseptic meningitis of unknown causes, were analyzed. Results were compared with the outcome of nested PCR for HSV-2 infection. Clinical parameters were analyzed in relation to CSF viral load. With real-time PCR, HSV-2 DNA was found in CSF from 80% (52/65) of patients with clinical HSV-2 meningitis compared to 72% (47/65) found by nested PCR. The sensitivity of real-time HSV-2 PCR was found to be 87% (33/38) in primary and 70% (19/27) in recurrent meningitis. The HSV-2 viral load was significantly higher in primary than in recurrent meningitis and correlated with the degree of inflammation. VZV DNA was detected in 2 of 45 samples (4.4%). Real-time PCR for the diagnosis of HSV-2 meningitis was evaluated in a large, clinically well-characterized sample of material and found to identify more cases than nested PCR in the group of patients with recurrent meningitis. Quantification of DNA enables further research of disease prognosis and treatment.

Transmission of cytomegalovirus to extremely preterm infants through breast milk.

AIM: To evaluate the rate and clinical expression of postnatal cytomegalovirus (CMV) infection transmitted through breast milk in extremely preterm infants. METHODS: Ten extremely preterm infants and their six mothers were included. Maternal CMV serology was determined. Breast milk samples and urine samples from the infants were screened for CMV. Symptoms and laboratory findings of CMV infected infants were documented. All infants received partly fresh and/or defrosted breast milk. RESULTS: CMV-DNA was found in breast milk in four of five CMV-seropositive mothers. Two infants were infected by CMV. They were the only infants fed with breast milk positive for viral culture. One infant developed hepatic affection concurrent with viral excretion in urine. This infant was later diagnosed with cystic fibrosis. CONCLUSION: This study supports that CMV transmission through breast milk can aggravate the clinical course in extremely preterm infants with preexisting hepatic conditions.

Identification of viral agents associated with diarrhea in young children during a winter season in Beijing, China.

BACKGROUND: Viral diarrhea remains a major cause of childhood morbidity and mortality worldwide. Although rotavirus was extensively studied in China, few comprehensive studies of all viral agents related to diarrhea in children have been conducted. OBJECTIVES: Our study was performed to investigate the role of enteric viruses in acute diarrhea in our country and to evaluate methods that could be used in routine diagnostics. STUDY DESIGN: One hundred stool samples were collected from children under 5 years of age seeking medical care for acute diarrhea during the winter season 2000/2001 in Beijing Children's Hospital. All specimens were initially screened microscopically for leucocytes/red blood cells. Samples with negative results were analyzed for virus presence using commercial EIAs and/or in-house RT-PCRs. RESULTS: At least one viral agent was found in 67% of the specimens. The frequency of rotavirus, astrovirus, norovirus and enteric adenovirus was 59%, 8%, 6% and 2%, respectively. Dual infections were found in 9.0% (6/67) of the positive samples. The results from rotavirus and astrovirus EIAs were concordant with those of rotavirus and astrovirus RT-PCRs. CONCLUSIONS: Enteric viruses play an important role in pediatric diarrhea during the winter season in China. A combination of microscopic examination of stool samples with specific EIA assays to detect virus antigen in stool specimens may be suitable for routine diagnostics.

Evaluation of the ReSSQ assay in relation to the COBAS AMPLICOR CMV MONITOR test and an in-house nested PCR method for detection of cytomegalovirus DNA.

The ReSSQ CMV assay is a novel commercially available kit for quantification of cytomegalovirus (CMV), based on real-time PCR with a peptide nucleic acid probe coupled with a single dye. In combination with the LightCycler, the ReSSQ CMV assay was evaluated with respect to specificity, PCR inhibition, linearity, reproducibility, and sensitivity. All nontested CMV materials were negative, and the assay was not inhibited by the use of different anticoagulants or other factors that may influence blood samples. The dynamic range was between 10 and 5 x 10(8) copies/PCR, and intra- and interassay variabilities were below 0.10 and 0.12 log10 standard deviations, respectively. Assay sensitivity was validated by analysis of 24 samples from a proficiency panel and by comparison to a nested in-house CMV PCR and the COBAS AMPLICOR CMV MONITOR test, using 159 clinical samples. Results from the proficiency panel were well in accordance with input values over the entire range of viral concentrations tested (50 to 31,250 copies/ml). The association between the ReSSQ CMV assay and the in-house PCR was in agreement in 90% of the clinical samples, and discordant results were found for all types of sample materials analyzed. The ReSSQ CMV and COBAS AMPLICOR assays showed no significant differences for samples containing >1,000 CMV copies/ml, but results differed to a greater extent at lower viral concentrations. The results demonstrate that the ReSSQ CMV assay is a CMV-specific, robust, and reproducible method and hence is well suited for routine use in clinical virology laboratories.

Molecular epidemiology of norovirus infections in Stockholm, Sweden, during the years 2000 to 2003: association of the GGIIb genetic cluster with infection in children.

The incidence of norovirus-associated gastroenteritis and the molecular epidemiology of norovirus strains were studied during three seasons (2000-2001, 2001-2002, and 2002-2003) among patients of all ages, mainly from the Stockholm region in Sweden. A total of 3,252 fecal samples were analyzed by reverse transcription-PCR. The incidences of norovirus infection among adults were 23, 26, and 30% during the three seasons studied and 18, 11, and 15% among children 0 to 15 years of age. During the first season, all norovirus strains detected by PCR were typed either by reverse line blot hybridization or nucleotide sequence analysis. During the two successive seasons, a total of 60 norovirus-positive strains from the beginning, peak, and end of the seasons were selected for nucleotide sequence analysis. We identified two dominant norovirus variants over the seasons: a new norovirus variant, recently described as the GGIIb genetic cluster, dominated among children during the first season, and during the following two seasons, a GGII-4 variant dominated. Our data suggest that norovirus infections are common, not only among adults, but also among children, and that some strains may predominantly affect children.

Respiratory virus infections in Stockholm during seven seasons: a retrospective study of laboratory diagnosis.

A retrospective analysis of the virological findings in all respiratory samples (7303) analysed at the laboratory of Karolinska Hospital between 1993 and 2000 was performed. The findings were studied according to age and seasonal variation, and the methods were evaluated. Most samples were from children. RSV was the dominant agent, found in 34% of all samples from children 0-1 y of age. Influenza A was found in 13% of samples from the age group 2-5 y. Influenza A dominated among adults and the elderly. RSV was found only in 2% of samples from patients 81 y or older. Adenovirus was found among children and adults, but not at all among the elderly. Both antigen detection and virus isolation were performed on 79% (5776) of the samples. For diagnosis of influenza A, virus isolation was more sensitive than immunofluorescence, but for diagnosis of RSV immunofluorescence was more sensitive than virus isolation. Thus, the analysis verified that influenza A is common not only among adults and the elderly, but also among small children. RSV was an uncommon finding among the elderly. Immunofluorescence is sensitive and rapid for the diagnosis of particularly RSV among small children and influenza in all age groups.

The occurrence of antibodies against Chlamydia species in patients with monoarthritis and chronic closed lock of the temporomandibular joint.

PURPOSE: The study goal was to investigate the occurrence of serum antibodies to Chlamydia trachomatis, Chlamydia pneumoniae, and Chlamydia psittaci in patients with temporomandibular joint (TMJ) monoarthritis or chronic closed lock and in control subjects. PATIENTS AND METHODS: An indirect microimmunofluorescence test for detecting antibodies against C trachomatis was used. Twenty-three patients (12 with monoarthritis and 11 with chronic closed lock) and 42 control subjects were evaluated. RESULTS: Six patients with monoarthritis, 5 patients with chronic closed lock of the TMJ, and 6 control individuals were considered to have had a past C trachomatis infection based on their immunoglobulin G titers. Corresponding groups for C pneumoniae investigation included 3 patients with monoarthritis, 4 patients with chronic closed lock, and 17 control subjects, and for C psittaci, 1 patient with monoarthritis, 2 patients with chronic closed lock, and 1 control subject. Statistically significant differences between patients and control subjects were found for C trachomatis only; monoarthritis versus control (P =.016), chronic closed lock versus control (P =.038), and all patients versus control (P =.007). Patients with monoarthritis did not differ from patients with chronic closed lock with regard to antibodies against C trachomatis. CONCLUSION: The occurrence of serum antibodies to C trachomatis was significantly higher in patients than in control subjects, but this occurrence did not correlate with severity of observed tissue changes. Nevertheless, an association may exist between the presence of C trachomatis and TMJ disease.