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We compared cycle thresholds from mpox skin lesions with other specimen sites and over time from onset of clinical signs among 104 patients in Sweden. Cycle thresholds differed by anatomic site. We identified 2 early mpox cases from anorectal swab specimens after skin samples were negative, indicating necessity of sampling multiple sites.

Abstract Background In order to estimate the prevalence and understand the spread of SARS-CoV-2 in Sweden, the Public Health Agency of Sweden, with support from the Swedish Armed Forces, conducted a series of point prevalence surveys between March and December 2020. Methods Sampling material and instructions on how to perform self-sampling of the upper respiratory tract were delivered to the homes of the participants. Samples were analysed by real-time PCR, and the participants completed questionnaires regarding symptoms. Findings The first survey in the Stockholm region in March 2020 included 707 participants and showed a SARS-CoV-2 prevalence of 2.5%. The following five surveys, performed on a national level, with between 2461 and 2983 participants, showed SARS-CoV-2 prevalences of 0.9% (April), 0.3% (May), 0.0% (August), 0.0% (September), and 0.7% (December). All positive cases who responded to questionnaires reported experiencing symptoms that occurred from 2 weeks before the date of sampling up to and including the date of sampling. Interpretation None of the individuals shown to be PCR-positive were asymptomatic at the time of sampling or in the 14 days prior to sampling. This is in contrast to many other surveys in which a substantial proportion of positive cases have been reported to be asymptomatic. Our surveys demonstrate a decreasing ratio between notified cases and the observed prevalence throughout the year, in line with increasing testing capacity and the consecutive inclusion of all symptomatic individuals in the case definition for testing.

A previously healthy male patient had detectable monkeypox virus DNA in saliva 76 days after laboratory confirmation of infection. A comprehensive characterization of viral kinetics and a detailed follow-up indicated a declining risk for transmission during the weeks after monkeypox symptoms appeared.

Abstract We evaluated the performance of 11 SARS-CoV-2 antibody tests using a reference set of heat-inactivated samples from 278 unexposed persons and 258 COVID-19 patients, some of whom contributed serial samples. The reference set included samples with a variation in SARS-CoV-2 IgG antibody titers, as determined by an in-house immunofluorescence assay (IFA). The five evaluated rapid diagnostic tests had a specificity of 99.0% and a sensitivity that ranged from 56.3 to 81.6% and decreased with low IFA IgG titers. The specificity was > 99% for five out of six platform-based tests, and when assessed using samples collected ≥ 22 days after symptom onset, two assays had a sensitivity of > 96%. These two assays also detected samples with low IFA titers more frequently than the other assays. In conclusion, the evaluated antibody tests showed a heterogeneity in their performances and only a few tests performed well with samples having low IFA IgG titers, an important aspect for diagnostics and epidemiological investigations.

SARS-CoV-2 infection is widely diagnosed by RT-PCR, but RNA extraction is a bottleneck for fast and cheap diagnosis. Here, the authors develop protocols to perform RT-PCR directly on heat-inactivated subject samples or samples lysed with readily available detergents and benchmark performance against 597 clinically diagnosed patient samples.

The emergence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) created an exceptional situation in which numerous laboratories in Europe simultaneously implemented SARS-CoV-2 diagnostics. These laboratories reported in February 2020 that commercial primer and probe batches for SARS-CoV-2 detection were contaminated with synthetic control material, causing delays of regional testing roll-out in various countries.

Torque teno virus (TTV) is an unenveloped, circular, single stranded DNA virus with a genome size of approximately 3.8 kb. Previous studies have demonstrated varying grades of association between TTV DNA levels and immune deficiencies related to age, chronic infections and cancer. Alzheimer's disease (AD) has been related to persistent viral infections such as HSV-1 and CMV, but it is not known whether TTV viral load could serve as a functional biomarker of cellular immunity in this setting. Therefore, the objective of this study was to investigate whether TTV infection and viral load is related to AD status, CMV immunity, systemic inflammation or HLA types connected to anti-viral immunity. A total of 50 AD subjects and 51 non-demented controls were included in the study. AD subjects were diagnosed according to NINCDS-ADRDA and DSM-IV criteria and neuroradiologic findings were consistent with the diagnosis. TTV viral load was analyzed in plasma samples using a quantitative real-time PCR. Using a cut-off for TTV status at 200 copies/ml, 88% (89/101) of the study subjects were classified as TTV positive. TTV viral load significantly increased with age (beta 0.049 per year, p

This study identified and characterized Brucella species in the informal milk chain in Uganda. A total of 324 cattle bulk milk samples were screened for the genus Brucella by real-time PCR with primers targeting the bcsp31 gene and further characterized by the omp25 gene. Of the samples tested, 6.5% were positive for Brucella species. In the omp25 phylogeny, the study sequences were found to form a separate clade within the branch containing B. abortus sequences. The study shows that informally marketed cattle milk in Uganda is a likely risk factor for human brucellosis and confirms that B. abortus is present in the cattle population. This information is important for potential future control measures, such as vaccination of cattle.

Coronaviruses (CoVs) are found in a wide variety of wild and domestic animals, and constitute a risk for zoonotic and emerging infectious disease. In poultry, the genetic diversity, evolution, distribution and taxonomy of some coronaviruses have been well described, but little is known about the features of CoVs in wild birds. In this study we screened 764 samples from 22 avian species of the orders Anseriformes and Charadriiformes in Sweden collected in 2006/2007 for CoV, with an overall CoV prevalence of 18.7%, which is higher than many other wild bird surveys. The highest prevalence was found in the diving ducks--mainly Greater Scaup (Aythya marila; 51.5%)--and the dabbling duck Mallard (Anas platyrhynchos; 19.2%). Sequences from two of the Greater Scaup CoV fell into an infrequently detected lineage, shared only with a Tufted Duck (Aythya fuligula) CoV. Coronavirus sequences from Mallards in this study were highly similar to CoV sequences from the sample species and location in 2011, suggesting long-term maintenance in this population. A single Black-headed Gull represented the only positive sample from the order Charadriiformes. Globally, Anas species represent the largest fraction of avian CoV sequences, and there seems to be no host species, geographical or temporal structure. To better understand the eitiology, epidemiology and ecology of these viruses more systematic surveillance of wild birds and subsequent sequencing of detected CoV is imperative.

Wild waterfowl is the natural reservoir of influenza A virus (IAV); hosted viruses are very variable and provide a source for genetic segments which can reassort with poultry or mammalian adapted IAVs to generate novel species crossing viruses. Additionally, wild waterfowl act as a reservoir for highly pathogenic IAVs. Exposure of wild birds to the antiviral drug oseltamivir may occur in the environment as its active metabolite can be released from sewage treatment plants to river water. Resistance to oseltamivir, or to other neuraminidase inhibitors (NAIs), in IAVs of wild waterfowl has not been extensively studied.In a previous in vivo Mallard experiment, an influenza A(H6N2) virus developed oseltamivir resistance by the R292K substitution in the neuraminidase (NA), when the birds were exposed to oseltamivir. In this study we tested if the resistance could be maintained in Mallards without drug exposure. Three variants of resistant H6N2/R292K virus were each propagated during 17 days in five successive pairs of naïve Mallards, while oseltamivir exposure was decreased and removed. Daily fecal samples were analyzed for viral presence, genotype and phenotype.Within three days without drug exposure no resistant viruses could be detected by NA sequencing, which was confirmed by functional NAI sensitivity testing. We conclude that this resistant N2 virus could not compete in fitness with wild type subpopulations without oseltamivir drug pressure, and thus has no potential to circulate among wild birds. The results of this study contrast to previous observations of drug induced resistance in an avian H1N1 virus, which was maintained also without drug exposure in Mallards. Experimental observations on persistence of NAI resistance in avian IAVs resemble NAI resistance seen in human IAVs, in which resistant N2 subtypes do not circulate, while N1 subtypes with permissive mutations can circulate without drug pressure. We speculate that the phylogenetic group N1 NAs may easier compensate for NAI resistance than group N2 NAs, though further studies are needed to confirm such conclusions.

Resistance to neuraminidase inhibitors (NAIs) is problematic as these drugs constitute the major treatment option for severe influenza. Extensive use of the NAI oseltamivir (Tamiflu®) results in up to 865 ng/L of its active metabolite oseltamivir carboxylate (OC) in river water. There one of the natural reservoirs of influenza A, dabbling ducks, can be exposed. We previously demonstrated that an influenza A(H1N1) virus in mallards (Anas platyrhynchos) exposed to 1 µg/L of OC developed oseltamivir resistance through the mutation H274Y (N2-numbering). In this study, we assessed the resistance development in an A(H6N2) virus, which belongs to the phylogenetic N2 group of neuraminidases with distinct functional and resistance characteristics. Mallards were infected with A(H6N2) while exposed to 120 ng/L, 1.2 µg/L or 12 µg/L of OC in their sole water source. After 4 days with 12 µg/L of OC exposure, the resistance mutation R292K emerged and then persisted. Drug sensitivity was decreased ≈13,000-fold for OC and ≈7.8-fold for zanamivir. Viral shedding was similar when comparing R292K and wild-type virus indicating sustained replication and transmission. Reduced neuraminidase activity and decrease in recovered virus after propagation in embryonated hen eggs was observed in R292K viruses. The initial, but not the later R292K isolates reverted to wild-type during egg-propagation, suggesting a stabilization of the mutation, possibly through additional mutations in the neuraminidase (D113N or D141N) or hemagglutinin (E216K). Our results indicate a risk for OC resistance development also in a N2 group influenza virus and that exposure to one NAI can result in a decreased sensitivity to other NAIs as well. If established in influenza viruses circulating among wild birds, the resistance could spread to humans via re-assortment or direct transmission. This could potentially cause an oseltamivir-resistant pandemic; a serious health concern as preparedness plans rely heavily on oseltamivir before vaccines can be mass-produced.

Oseltamivir (Tamiflu®) is the most widely used drug against influenza infections and is extensively stockpiled worldwide as part of pandemic preparedness plans. However, resistance is a growing problem and in 2008-2009, seasonal human influenza A/H1N1 virus strains in most parts of the world carried the mutation H274Y in the neuraminidase gene which causes resistance to the drug. The active metabolite of oseltamivir, oseltamivir carboxylate (OC), is poorly degraded in sewage treatment plants and surface water and has been detected in aquatic environments where the natural influenza reservoir, dabbling ducks, can be exposed to the substance. To assess if resistance can develop under these circumstances, we infected mallards with influenza A/H1N1 virus and exposed the birds to 80 ng/L, 1 µg/L and 80 µg/L of OC through their sole water source. By sequencing the neuraminidase gene from fecal samples, we found that H274Y occurred at 1 µg/L of OC and rapidly dominated the viral population at 80 µg/L. IC₅₀ for OC was increased from 2-4 nM in wild-type viruses to 400-700 nM in H274Y mutants as measured by a neuraminidase inhibition assay. This is consistent with the decrease in sensitivity to OC that has been noted among human clinical isolates carrying H274Y. Environmental OC levels have been measured to 58-293 ng/L during seasonal outbreaks and are expected to reach µg/L-levels during pandemics. Thus, resistance could be induced in influenza viruses circulating among wild ducks. As influenza viruses can cross species barriers, oseltamivir resistance could spread to human-adapted strains with pandemic potential disabling oseltamivir, a cornerstone in pandemic preparedness planning. We propose surveillance in wild birds as a measure to understand the resistance situation in nature and to monitor it over time. Strategies to lower environmental levels of OC include improved sewage treatment and, more importantly, a prudent use of antivirals.

BACKGROUND: The recent report of gammaretroviruses of probable murine origin in humans, called xenotropic murine retrovirus related virus (XMRV) and human murine leukemia virus related virus (HMRV), necessitated a bioinformatic search for this virus in genomes of the mouse and other vertebrates, and by PCR in humans. RESULTS: Three major groups of murine endogenous gammaretroviruses were identified. The third group encompassed both exogenous and endogenous Murine Leukemia Viruses (MLVs), and most XMRV/HMRV sequences reported from patients suffering from myalgic encephalomyelitis/chronic fatigue syndrome (ME/CFS). Two sensitive real-time PCRs for this group were developed. The predicted and observed amplification range for these and three published XMRV/HMRV PCRs demonstrated conspicuous differences between some of them, partly explainable by a recombinatorial origin of XMRV. Three reverse transcription real-time PCRs (RTQPCRs), directed against conserved and not overlapping stretches of env, gag and integrase (INT) sequences of XMRV/HMRV were used on human samples. White blood cells from 78 patients suffering from ME/CFS, of which 30 patients also fulfilled the diagnostic criteria for fibromyalgia (ME/CFS/FM) and in 7 patients with fibromyalgia (FM) only, all from the Gothenburg area of Sweden. As controls we analyzed 168 sera from Uppsala blood donors. We controlled for presence and amplifiability of nucleic acid and for mouse DNA contamination. To score as positive, a sample had to react with several of the XMRV/HMRV PCRs. None of the samples gave PCR reactions which fulfilled the positivity criteria. CONCLUSIONS: XMRV/HMRV like proviruses occur in the third murine gammaretrovirus group, characterized here. PCRs developed by us, and others, approximately cover this group, except for the INT RTQPCR, which is rather strictly XMRV specific. Using such PCRs, XMRV/HMRV could not be detected in PBMC and plasma samples from Swedish patients suffering from ME/CFS/FM, and in sera from Swedish blood donors.

Coronaviruses (CoVs) can cause mild to severe disease in humans and animals, their host range and environmental spread seem to have been largely underestimated, and they are currently being investigated for their potential medical relevance. Infectious bronchitis virus (IBV) belongs to gamma-coronaviruses and causes a costly respiratory viral disease in chickens. The role of wild birds in the epidemiology of IBV is poorly understood. In the present study, we examined 1,002 cloacal and faecal samples collected from 26 wild bird species in the Beringia area for the presence of CoVs, and then we performed statistical and phylogenetic analyses. We detected diverse CoVs by RT-PCR in wild birds in the Beringia area. Sequence analysis showed that the detected viruses are gamma-coronaviruses related to IBV. These findings suggest that wild birds are able to carry gamma-coronaviruses asymptomatically. We concluded that CoVs are widespread among wild birds in Beringia, and their geographic spread and frequency is higher than previously realised. Thus, Avian CoV can be efficiently disseminated over large distances and could be a genetic reservoir for future emerging pathogenic CoVs. Considering the great animal health and economic impact of IBV as well as the recent emergence of novel coronaviruses such as SARS-coronavirus, it is important to investigate the role of wildlife reservoirs in CoV infection biology and epidemiology.

Objective: The aim of this study was to compare the safety and efficacy of aspirin 162 mg to the standard recommended dose of 81 mg for preeclampsia prevention., Study Design: A retrospective cohort study of patients at risk for preeclampsia who delivered between January 2013 and December 2020 at Henry Ford Health was performed. Patients were divided into three groups: a no aspirin group, a group treated under an 81 mg aspirin preeclampsia prophylaxis protocol, and a group treated under a 162 mg protocol. Univariate and multivariable logistic regression analyses compared rates of preeclampsia and secondary outcomes between groups. Clinical side effects traditionally associated with aspirin use were also assessed., Results: Of 3,597 patients, 2,266 (63%) were in the no aspirin group, 944 (26%) were in the 81 mg group, and 387 (11%) were in the 162 mg group. The rate of preeclampsia was significantly lower in the 162 mg group (10.1%, odds ratio, 0.68; 95% confidence interval, 0.46-0.99) compared with the 81 mg group (14.2%). The rate of preeclampsia was identical in the no aspirin and 81 mg groups. The rate for postpartum hemorrhage, postpartum hematoma, and intraventricular hemorrhage of the newborn were not significantly different between patients in the 162 and 81 mg groups., Conclusion: We observed a significantly lower rate of preeclampsia in high-risk patients who were treated with the 162 mg dose of aspirin for preeclampsia prophylaxis, and bleeding complications were not seen with the higher dose. Our study suggests that aspirin 162 mg may be considered for prophylaxis in patients at high risk for preeclampsia., Key Points: · Aspirin 81 mg is currently standard for preeclampsia prophylaxis.. · Preeclampsia rate is significantly lower among high-risk patients taking aspirin 162 mg compared with 81 mg.. · Bleeding complications are not increased among those taking aspirin 162 mg.., Competing Interests: None declared., (Thieme. All rights reserved.)

Objective: To evaluate provider adherence to aspirin prophylaxis prescription guidelines for patients at risk., Study Design: A retrospective chart review was performed at Henry Ford Health (HFH) between October 2015 and December 2020. In October 2015, low-dose aspirin was recommended for women who met high risk criteria for preeclampsia at HFH; in February 2019, aspirin recommendation expanded to include women who met either moderate or high-risk criteria. A total of 46,016 pregnancies occurred between Oct 2015 and Dec 2020 of which 15,167 (33.0%) met high and moderate risk criteria., Results: From the population at risk, 1,255 (8.3%) had a history of preeclampsia, 2,534 (16.7%) had a history of chronic hypertension, 1,418 (9.3%) had a history of diabetes, 7,470 (49.3%) were nulliparous, 4,038 (26.6%) were 35 years of age or older, 6,395 (42.2%) had a body mass index greater than 30 kg/m2, and 8,174 (54.5%) were African Americans. Only 630 out of 3,584 (17.6%) of women meeting the high-risk criteria for preeclampsia between Oct 2015 and Jan 2019 received low-dose aspirin and only 891 out of 5,874 (15.2%) of women meeting the high or moderate risk criteria for preeclampsia between Feb 2019 and Dec 2020 received low-dose aspirin prophylaxis., Conclusion: Adherence to aspirin prophylaxis guidelines was low. Most urban healthcare systems serve diverse, high-risk populations with multiple comorbidities rendering many women at risk for preeclampsia. Educational efforts to improve provider knowledge regarding this important preventative measure are indicated. Recommendation for implementing universal aspirin in such high-risk populations should also be considered., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Published by Elsevier B.V.)

False-positive human immunodeficiency virus (HIV) test results are rare but have been documented in the setting of certain underlying conditions such as Epstein-Barr virus, metastatic cancer, and certain autoimmune conditions. A retrospective cohort study in a large hospital system was conducted to compare the occurrence of false-positive HIV fourth-generation test results before and after the coronavirus disease 2019 (COVID-19) pandemic in a population of pregnant patients (N=44,187; 22,073 pre-COVID and 22,114 during COVID). The COVID cohort had a significantly higher frequency of false-positive HIV test results compared with the pre-COVID cohort (0.381 vs 0.676, P =.002). Within the COVID cohort, 25% of patients had a positive polymerase chain reaction test result for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) preceding their false-positive HIV test results. When this subgroup was excluded, the difference in frequency of false-positive HIV test results between the cohorts was no longer significant (0.381 vs 0.507, P =.348). Our findings suggest that SARS-CoV-2 seropositivity was associated with an increased frequency of false-positive HIV test results in the pregnant population., Competing Interests: Financial Disclosure The authors did not report any potential conflicts of interest., (Copyright © 2023 by the American College of Obstetricians and Gynecologists. Published by Wolters Kluwer Health, Inc. All rights reserved.)

Introduction: Coronavirus disease 2019 (COVID-19) results in similar clinical characteristics as bacterial respiratory tract infections and can potentially lead to antibiotic overuse. This study aimed to determine the changes in hospital antimicrobial usage before and during the COVID-19 pandemic., Methodology: We compared antimicrobial consumption data for 2019 and 2020. Inpatient antibiotic consumption was determined and expressed as a defined daily dose (DDD) per 100 occupied bed days, following the World Health Organization (WHO) methods. The WHO Access, Watch, and Reserve (AWaRe) classification was used., Results: The total antimicrobial consumption in 2020 increased by 16.3% compared to consumption in 2019. In 2020, there was a reduction in fourth-generation cephalosporins (-30%), third-generation cephalosporins (-29%), and combinations of penicillins (-23%). In contrast, antibiotics that were consumed more during 2020 compared with 2019 included linezolid (374%), vancomycin (66.6%), and carbapenem (7%). Linezolid is the only antibiotic from the Reserve group on the hospital's formulary. Antibiotic usage from the Access group was reduced by 17%, while antibiotic usage from the Watch group and the Reserve group was increased by 3% and 374%, respectively., Conclusions: The findings show a significant shift in antibiotic usage from the Access group to the Watch and Reserve groups. The Watch and Reserve groups are known to be associated with increased resistance to antibiotics. Therefore, antimicrobial stewardship should be increased and maintained during the pandemic to ensure appropriate antibiotic use., Competing Interests: No Conflict of Interest is declared, (Copyright (c) 2022 Raghda RS Hussein, Al Shaimaa Ibrahim Rabie, Maryam Bin Shaman, Ahmed Hassan Shaaban, Alzhraa M Fahmy, Mahmoud R Sofy, Elizabeth A Lattyak, Ahmed Abuelhana, Ibrahim A Naguib, Ahmed M Ashour, Mamoon A Aldeyab.)

The present study aimed to survey medical, nursing and pharmacy students' knowledge, attitude and practice regarding antimicrobial use and resistance. Additionally, the study assessed the teaching and assessment activities received regarding antibiotic use. A cross sectional online survey was distributed to undergraduate students currently in clinical studies in their degree program. A total of 716 medicine, nursing and pharmacy undergraduate students were included. Respondents scored more than 76% on knowledge on effective use, unnecessary use and associated side effects of antibiotics, and 65.2% regarding knowledge on the spread of antibiotic resistance. Some participants (21.0%) agreed or strongly agreed that there has been good promotion of prudent antimicrobial use. Students were aware (13.1%), unaware (29.1%), or unsure (57.8%) that there is a national action plan relating to antimicrobial resistance. A total of 62.8% of the respondents strongly agreed or agreed that they have a key role in helping control antibiotic resistance. Participants reported that they require more information about resistance to antibiotics (53.9%), medical conditions for which antibiotics are used (51.7%) and how to use antibiotics (51.0%). Discussion of clinical cases and vignettes and small group teaching were reported as very useful or useful teaching strategies (79.9% and 74.2%, respectively). The findings from this study determined the current situation in relation to education on prudent antimicrobial use for undergraduates and highlighted areas for informing better curriculum design.

Objective: To examine the rate of recurrence for elevated isolated maternal serum alpha-fetoprotein (MSAFP) and its associated adverse outcomes during a subsequent pregnancy. Materials and Methods: A retrospective cohort study of pregnant multiparous women who had elevated MSAFP levels during an initial and a subsequent pregnancy between 1994 and 2020. Results: Twenty-seven out of 344 (7.8%) women with elevated MSAFP had recurrent elevated MSAFP in a subsequent pregnancy. Four women were excluded due to missing data. Of the 23 women included, 5 (22%) had fetal growth restriction (FGR), 2 (9%) had pre-eclampsia, 9 (35%) had preterm births, and 2 (9%) had fetal death/miscarriage in their subsequent pregnancy. Looking at individual outcomes, 60% of women had recurrence of preterm labor, 33% had recurrence of fetal death, and 25% had recurrence of FGR. Conclusion: Women with elevated MSAFP levels during an initial pregnancy should be informed during preconception counseling about their risk of recurring elevated MSAFP and its associated adverse outcomes risks.

Background Microbial invasion of the amniotic cavity resulting in intra-amniotic infection is associated with obstetrical complications such as preterm labor with intact or ruptured membranes, cervical insufficiency, as well as clinical and histological chorioamnionitis. The most widely accepted pathway for intra-amniotic infection is the ascension of microorganisms from the lower genital tract. However, hematogenous dissemination of microorganisms from the oral cavity or intestine, retrograde seeding from the peritoneal cavity through the fallopian tubes, and introduction through invasive medical procedures have also been suggested as potential pathways for intra-amniotic infection. The primary reason that an ascending pathway is viewed as most common is that the microorganisms most often detected in the amniotic fluid are those that are typical inhabitants of the vagina. However, thus far, no studies have shown that microorganisms in the amniotic cavity are simultaneously present in the vagina of the woman from which they were isolated. The objective of the study was to determine the frequency with which microorganisms isolated from women with intra-amniotic infection are also present in the lower genital tract. Methods This was a cross-sectional study of women with intra-amniotic infection with intact membranes. Intra-amniotic infection was defined as a positive culture and elevated concentrations of interleukin-6 (IL-6) (>2.6 ng/mL) in amniotic fluid and/or acute histologic chorioamnionitis and funisitis. Microorganisms isolated from bacterial cultures of amniotic fluid were taxonomically identified through matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF) and 16S ribosomal RNA (rRNA) gene sequencing. Vaginal swabs were obtained at the time of amniocentesis for the identification of microorganisms in the lower genital tract. The overall bacterial profiles of amniotic fluids and vaginal swabs were characterized through 16S rRNA gene sequencing. The bacterial profiles of vaginal swabs were interrogated for the presence of bacteria cultured from amniotic fluid and for the presence of prominent (>1% average relative abundance) operational taxonomic units (OTUs) within the overall 16S rRNA gene bacterial profiles of amniotic fluid. Results (1) A total of 75% (6/8) of women had bacteria cultured from their amniotic fluid that are typical residents of the vaginal ecosystem. (2) A total of 62.5% (5/8) of women with bacteria cultured from their amniotic fluid also had these bacteria present in their vagina. (3) The microorganisms cultured from amniotic fluid and also detected in the vagina were Ureaplasma urealyticum, Escherichia coli, and Streptococcus agalactiae. (4) 16S rRNA gene sequencing revealed that the amniotic fluid of women with intra-amniotic infection had bacterial profiles dominated by Sneathia, Ureaplasma, Prevotella, Lactobacillus, Escherichia, Gardnerella, Peptostreptococcus, Peptoniphilus, and Streptococcus, many of which had not been cultured from the amniotic fluid samples. (5) Seventy percent (7/10) of the prominent (>1% average relative abundance) OTUs found in amniotic fluid were also prominent in the vagina. Conclusion The majority of women with intra-amniotic infection had bacteria cultured from their amniotic fluid that were typical vaginal commensals, and these bacteria were detected within the vagina at the time of amniocentesis. Molecular microbiological interrogation of amniotic fluid from women with intra-amniotic infection revealed that the bacterial profiles of amniotic fluid were largely consistent with those of the vagina. These findings indicate that ascension from the lower genital tract is the primary pathway for intra-amniotic infection.

Objective: Neutrophil extracellular traps (NETs) control microbial infections through their antimicrobial activities attributed to DNA, histones, granules, and cytoplasmic proteins (eg, elastase). Intra-amniotic infection is characterized by the influx of neutrophils into the amniotic cavity; therefore, the aim of this study was to determine whether amniotic fluid neutrophils form NETs in this inflammatory process., Methods: Amniotic fluid samples from women with intra-amniotic infection (n = 15) were stained for bacteria detection using fluorescent dyes. Amniotic fluid neutrophils were purified by filtration. As controls, neutrophils from maternal blood samples (n = 3) were isolated by density gradients. Isolated neutrophils were plated onto glass cover slips for culture with and without 100 nM of phorbol-12-myristate-13-acetate (PMA). NET formation was assessed by 4',6-diamidino-2-phenylindole (DAPI) staining and scanning electron microscopy. Different stages of NET formation were visualized using antibodies against elastase and histone H3, in combination with DAPI staining, by confocal microscopy. Finally, maternal or neonatal neutrophils were added to amniotic fluid samples from women without intra-amniotic infection (n = 4), and NET formation was evaluated by DAPI staining., Results: (1) NETs were present in the amniotic fluid of women with intra-amniotic infection; (2) all of the amniotic fluid samples had detectable live and dead bacteria associated with the presence of NETs; (3) in contrast to neutrophils from the maternal circulation, amniotic fluid neutrophils did not require PMA stimulation to form NETs; (4) different stages of NET formation were observed by co-localizing elastase, histone H3, and DNA in amniotic fluid neutrophils; and (5) neither maternal nor neonatal neutrophils form NETs in the amniotic fluid of women without intra-amniotic infection., Conclusion: NETs are detectable in the amniotic fluid of women with intra-amniotic infection.

Objectives: 1) To characterize the cellular composition of the amniotic fluid of patients diagnosed with clinical chorioamnionitis at term, as a function of the presence or absence of microorganisms determined by cultivation techniques, and 2) to characterize the cytokine production by white blood cells present in the amniotic fluid using flow cytometry-based techniques., Materials and Methods: Amniotic fluid samples from 20 women who had the diagnosis of clinical chorioamnionitis at term were analyzed using cultivation techniques (for aerobic and anaerobic bacteria as well as genital Mycoplasmas). Amniotic fluid IL-6 concentrations were determined by an enzyme-linked immunosorbent assay. Amniotic fluid leukocytes were visualized by using hematoxylin and eosin staining and immunofluorescence. Immunophenotyping of surface markers and cytokines was performed in amniotic fluid leukocytes using flow cytometry., Results: 1) Neutrophils (CD45+CD15+ cells) were the most common leukocyte subset found in the amniotic fluid, followed by monocytes (CD45+CD14+ cells); other white blood cells (such as lymphocytes and natural killer cells) were scarce in the amniotic fluid; 2) the absolute counts of neutrophils and monocytes were significantly higher in patients with microorganisms found in the amniotic fluid than in those without detectable microorganisms, using cultivation techniques; 3) there was a significant correlation between the absolute counts of neutrophils and monocytes determined by flow cytometry (Spearman's correlation=0.97; P<0.001); 4) there was a significant correlation between the absolute white blood cell count determined with a hemocytometer chamber and by flow cytometric analysis (Spearman's correlation=0.88; P<0.001); and 5) the profile of cytokine expression differed between monocytes and neutrophils; while neutrophils predominantly produced TNF-α and MIP-1β, monocytes expressed higher levels of IL-1β and IL-1α., Conclusion: Flow cytometry analysis of the amniotic fluid of patients with intra-amniotic infection and clinical chorioamnionitis at term demonstrated that neutrophils and monocytes are the most common cells participating in the inflammatory process. We have characterized, for the first time, the differential cytokine expression by these cells in this important complication of pregnancy.

Objective: Acute atherosis is a lesion of the spiral arteries characterized by fibrinoid necrosis of the vessel wall, an accumulation of fat-containing macrophages, and a mononuclear perivascular infiltrate, which can be found in patients with preeclampsia, fetal death, small-for-gestational age, spontaneous preterm labor/premature prelabor rupture of membrane, and spontaneous mid-trimester abortion. This lesion is thought to decrease blood flow to the intervillous space which may lead to other vascular lesions of the placenta. The objective of this study was to test whether there is an association between acute atherosis and placental lesions that are consistent with maternal vascular underperfusion (MVU), amniotic fluid infection (AFI), fetal vascular thrombo-occlusive disease (FVTOD) or chronic inflammation., Material and Methods: A retrospective cohort study of pregnant women who delivered between July 1998 and July 2014 at Hutzel Women's Hospital/Detroit Medical Center was conducted examine 16 457 placentas. The frequency of placenta lesions (diagnosed using the criteria of the Perinatal Section of the Society for Pediatric Pathology) was compared between pregnancies with and without acute atherosis., Results: Among 16 457 women who were enrolled, 10.2% (1671/16 457) were excluded, leaving 14 786 women who contributed data for analysis. Among them, the prevalence of acute atherosis was 2.2% (326/14 786). Women with acute atherosis were more than six times as likely as those without to have placental lesions consistent with maternal underperfusion (adjusted odds ratio - aOR: 6.7; 95% CI 5.2-8.6). To a lesser degree, acute atherosis was also associated with greater risks of having either lesions consistent with FVTOD (aOR 1.7; 95% CI 1.2-2.3) or chronic chorioamnionitis (aOR 1.9; 95% CI 1.3-3), but not with other chronic inflammatory lesions, after adjusting for gestational age at delivery. In contrast, women with acute atherosis were 60% less likely to have lesions consistent with AFI, adjusting for gestational age at delivery (aOR 0.4; 95% CI 0.3-0.5)., Conclusions: Acute atherosis is associated with increased risks of having placental lesions consistent with MVU, and to a lesser extent, chronic chorioamnionitis and those consistent with FVTOD.

Objective: The objectives of this study were to: (1) determine the amniotic fluid (AF) microbiology of patients with preterm prelabor rupture of membranes (PROM); and (2) examine the relationship between intra-amniotic inflammation with and without microorganisms (sterile inflammation) and adverse pregnancy outcomes in patients with preterm PROM., Methods: AF samples obtained from 59 women with preterm PROM were analyzed using cultivation techniques (for aerobic and anaerobic bacteria as well as genital mycoplasmas) and with broad-range polymerase chain reaction coupled with electrospray ionization mass spectrometry (PCR/ESI-MS). AF concentration of interleukin-6 (IL-6) was determined using ELISA. Results of both tests were correlated with AF IL-6 concentrations and the occurrence of adverse obstetrical/perinatal outcomes., Results: (1) PCR/ESI-MS, AF culture, and the combination of these two tests each identified microorganisms in 36% (21/59), 24% (14/59) and 41% (24/59) of women with preterm PROM, respectively; (2) the most frequent microorganisms found in the amniotic cavity were Sneathia species and Ureaplasma urealyticum; (3) the frequency of microbial-associated and sterile intra-amniotic inflammation was overall similar [ 29% (17/59)]: however, the prevalence of each differed according to the gestational age when PROM occurred; (4) the earlier the gestational age at preterm PROM, the higher the frequency of both microbial-associated and sterile intra-amniotic inflammation; (5) the intensity of the intra-amniotic inflammatory response against microorganisms is stronger when preterm PROM occurs early in pregnancy; and (6) the frequency of acute placental inflammation (histologic chorioamnionitis and/or funisitis) was significantly higher in patients with microbial-associated intra-amniotic inflammation than in those without intra-amniotic inflammation [93.3% (14/15) versus 38% (6/16); p = 0.001]., Conclusions: (1) The frequency of microorganisms in preterm PROM is 40% using both cultivation techniques and PCR/ESI-MS; (2) PCR/ESI-MS identified microorganisms in the AF of 50% more women with preterm PROM than AF culture; and (3) sterile intra-amniotic inflammation was present in 29% of these patients, and it was as or more common than microbial-associated intra-amniotic inflammation among those presenting after, but not before, 24 weeks of gestation.

Objective: Acute atherosis is characterized by subendothelial lipid-filled foam cells, fibrinoid necrosis and perivascular lymphocytic infiltration. This lesion is generally confined to non-transformed spiral arteries and is frequently observed in patients with preeclampsia. However, the frequency of acute atherosis in the great obstetrical syndromes is unknown. The purpose of this study was to determine the frequency and topographic distribution of acute atherosis in placentas and placental bed biopsy samples obtained from women with normal pregnancy and those affected by the "great obstetrical syndromes". We also examined the relationship between acute atherosis and pregnancy outcome in patients with preeclampsia., Material and Methods: A retrospective cohort study of pregnant women who delivered between July 1998 and July 2014 at Hutzel Women's Hospital/Detroit Medical Center was conducted to examine 16, 345 placentas. Patients were classified into the following groups: (1) uncomplicated pregnancy; (2) spontaneous preterm labor (sPTL) and preterm prelabor rupture of membranes (PPROM); (3) preeclampsia; (4) gestational hypertension; (5) small-for-gestational age (SGA); (6) chronic hypertension; (5) fetal death; (6) spontaneous abortion and (7) others. A subset of patients had placental bed biopsy. The incidence of acute atherosis was compared among the different groups., Results: (1) The prevalence of acute atherosis in uncomplicated pregnancies was 0.4% (29/6961) based upon examination of nearly 7000 placentas; (2) the frequency of acute atherosis was 10.2% (181/1779) in preeclampsia, 9% (26/292) in fetal death, 2.5% (3/120) in midtrimester spontaneous abortion, 1.7% (22/1,298) in SGA neonates and 1.2% (23/1,841) in sPTL and PPROM; (3) among patients with preeclampsia, those with acute atherosis than in those without the lesion had significantly more severe disease, earlier onset, and a greater frequency of SGA neonates (p < 0.05 all) and (4) the lesion was more frequently observed in the decidua (parietalis or basalis) than in the decidual segment of the spiral arteries in patients with placental bed biopsies., Conclusions: Acute atherosis is rare in normal pregnancy, and occurs more frequently in patients with pregnancy complications, including preeclampsia, sPTL, preterm PROM, midtrimester spontaneous abortion, fetal death and SGA.

Introduction: The objectives of this study were: 1) to determine the amniotic fluid (AF) microbiology of patients with the diagnosis of clinical chorioamnionitis at term using both cultivation and molecular techniques; and 2) to examine the relationship between intra-amniotic inflammation with and without microorganisms and placental lesions consistent with acute AF infection., Methods: The AF samples obtained by transabdominal amniocentesis from 46 women with clinical signs of chorioamnionitis at term were analyzed using cultivation techniques (for aerobic and anerobic bacteria as well as genital mycoplasmas) and broad-range polymerase chain reaction (PCR) coupled with electrospray ionization mass spectrometry (PCR/ESI-MS). The frequency of microbial invasion of the amniotic cavity (MIAC), intra-amniotic inflammation [defined as an AF interleukin 6 (IL-6) concentration ≥2.6 ng/mL], and placental lesions consistent with acute AF infection (acute histologic chorioamnionitis and/or acute funisitis) were examined according to the results of AF cultivation and PCR/ESI-MS as well as AF IL-6 concentrations., Results: 1) Culture identified bacteria in AF from 46% (21/46) of the participants, whereas PCR/ESI-MS was positive for microorganisms in 59% (27/46) – combining these two tests, microorganisms were detected in 61% (28/46) of patients with clinical chorioamnionitis at term. Eight patients had discordant test results; one had a positive culture and negative PCR/ESI-MS result, whereas seven patients had positive PCR/ESI-MS results and negative cultures. 2) Ureaplasma urealyticum (n=8) and Gardnerella vaginalis (n=10) were the microorganisms most frequently identified by cultivation and PCR/ESI-MS, respectively. 3) When combining the results of AF culture, PCR/ESI-MS and AF IL-6 concentrations, 15% (7/46) of patients did not have intra-amniotic inflammation or infection, 6.5% (3/46) had only MIAC, 54% (25/46) had microbial-associated intra-amniotic inflammation, and 24% (11/46) had intra-amniotic inflammation without detectable microorganisms. 4) Placental lesions consistent with acute AF infection were significantly more frequent in patients with microbial-associated intra-amniotic inflammation than in those without intra-amniotic inflammation [70.8% (17/24) vs. 28.6% (2/7); P=0.04]., Conclusion: Microorganisms in the AF were identified in 61% of patients with clinical chorioamnionitis at term; 54% had microbial-associated intra-amniotic inflammation, whereas 24% had intra-amniotic inflammation without detectable microorganisms.

Restless legs syndrome (RLS) and nausea and vomiting of pregnancy (NVP) are both common maternal conditions affecting quality of life. Gabapentin is currently FDA-approved for treating RLS and preliminary results have shown it may be effective for treating the most severe form of NVP, hyperemesis gravidarum (HG). Because NVP and HG symptoms peak early in pregnancy, the potential teratogenicity of gabapentin needs to be considered. We reviewed published pregnancy registries and cohorts for pregnancy outcomes associated with maternal gabapentin use. Gabapentin exposures from 5 pregnancy registries, 1 HG pilot study and 2 additional cases were reviewed. Among 294 first trimester gabapentin-monotherapy exposures, there were 5 major congenital malformations (MCMs) reported (1.7%), which favorably compares to the MCM rate in the general population (1.6-2.2%). Two of the registries reported maternal gabapentin use among 261 singleton pregnancies to be associated with roughly equivalent rates of premature birth, birth weight after correction for gestational age at delivery and maternal hypertension/eclampsia as those that have been reported in the general population. These data support the safety of gabapentin use in pregnancy; however, the number of exposures to date is still small. If future pregnancy registry data confirm this positive safety profile, gabapentin therapy would likely be a safe and effective treatment for RLS during pregnancy. Controlled, clinical trials are needed to assess gabapentin's effectiveness for HG., (Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.)

The authors describe various patterns of F-18 fluorodeoxyglucose (FDG) accumulation in the diaphragm and crura. They present 6 patients in whom incidental diaphragmatic and crural uptake of FDG was observed during positron emission tomography (PET). Hyperventilation is thought to be the potential underlying mechanism of this condition.