Marrying Step Feed with Secondary Clarifier Improvements to Significantly Increase Peak Wet Weather Treatment Capacity: An Integrated Methodology.

The need to increase the peak wet weather secondary treatment capacity of the City of Akron, Ohio, Water Reclamation Facility (WRF) provided the opportunity to test an integrated methodology for maximizing the peak wet weather secondary treatment capacity of activated sludge systems. An initial investigation, consisting of process modeling of the secondary treatment system and computational fluid dynamics (CFD) analysis of the existing relatively shallow secondary clarifiers (3.3 and 3.7 m sidewater depth in 30.5 m diameter units), indicated that a significant increase in capacity from 416 000 to 684 000 m3/d or more was possible by adding step feed capabilities to the existing bioreactors and upgrading the existing secondary clarifiers. One of the six treatment units at the WRF was modified, and an extensive 2-year testing program was conducted to determine the total peak wet weather secondary treatment capacity achievable. The results demonstrated that a peak wet weather secondary treatment capacity approaching 974 000 m3/d is possible as long as secondary clarifier solids and hydraulic loadings could be separately controlled using the step feed capability provided. Excellent sludge settling characteristics are routinely experienced at the City of Akron WRF, raising concerns that the identified peak wet weather secondary treatment capacity could not be maintained should sludge settling characteristics deteriorate for some reason. Computational fluid dynamics analysis indicated that the impact of the deterioration of sludge settling characteristics could be mitigated and the identified peak wet weather secondary treatment capacity maintained by further use of the step feed capability provided to further reduce secondary clarifier solids loading rates at the identified high surface overflow rates. The results also demonstrated that effluent limits not only for total suspended solids (TSS) and five-day carbonaceous biochemical oxygen demand (cBOD5) could be maintained, but also for ammonia-nitrogen and total phosphorous (TP). Although hydraulic limitations in other parts of the WRP prevent this full capacity to be realized, the City is proceeding to implement the modifications identified using this integrated methodology.

Characterizing shallow secondary clarifier performance where conventional flux theory over-estimates allowable solids loading rate.

The performance characteristics of relatively shallow (3.3 and 3.7 m sidewater depth in 30.5 m diameter) activated sludge secondary clarifiers were extensively evaluated during a 2-year testing program at the City of Akron Water Reclamation Facility (WRF), Ohio, USA. Testing included hydraulic and solids loading stress tests, and measurement of sludge characteristics (zone settling velocity (ZSV), dispersed and flocculated total suspended solids), and the results were used to calibrate computational fluid dynamic (CFD) models of the various clarifiers tested. The results demonstrated that good performance could be sustained at surface overflow rates in excess of 3 m/h, as long as the clarifier influent mixed liquor suspended solids (MLSS) concentration was controlled to below critical values. The limiting solids loading rate (SLR) was significantly lower than the value predicted by conventional solids flux analysis based on the measured ZSV/MLSS relationship. CFD analysis suggested that this resulted because mixed liquor entering the clarifier was being directed into the settled sludge blanket, diluting it and also creating a 'thin' concentration sludge blanket that overlays the thicker concentration sludge blanket typically expected. These results indicate the need to determine the allowable SLR for shallow clarifiers using approaches other than traditional solids flux analysis. A combination of actual testing and CFD analyses are demonstrated here to be effective in doing so.

Real-time qualitative PCR for 57 human adenovirus types from multiple specimen sources.

Human adenoviruses (HAdVs) are ubiquitous double-stranded DNA viruses that cause a wide array of diseases in humans including pharyngitis, pneumonia, gastroenteritis, hemorrhagic cystitis, and keratoconjunctivitis. They also cause life-threatening opportunistic infections in immunocompromised individuals and are responsible for outbreaks in certain populations. Diagnosis is traditionally by cell culture or antigen detection methods. However, some HAdVs can take up to 4 weeks to isolate, and diarrheagenic types 40 and 41 will not grow in routine cell culture. Therefore, the goal of this study was to design a rapid, real-time PCR assay to detect all known 57 HAdV types from multiple different specimen sources. Primers and fluorescence resonance energy transfer hybridization probes were designed to target a 185-bp region of the penton base gene of HAdV. The analytical sensitivity was determined to be 10 copies/µl for HAdV types showing exact primer/probe homology in specimen matrix. Using whole-virus strains, the analytical sensitivity for representative HAdV types ranged from 10(-1) to 10(3) 50% tissue culture infective dose (TCID(50))/ml. The assay demonstrated 100% sensitivity and 99% specificity. This real-time PCR assay provides a rapid method for the detection of all 57 known HAdV types from respiratory specimens, blood, stool, urine, and ocular swabs.

Quantitative real-time polymerase chain reaction for evaluating DNAemia due to cytomegalovirus, Epstein-Barr virus, and BK virus in solid-organ transplant recipients.

Testing for cytomegalovirus-, Epstein-Barr virus-, and BK virus-specific gene targets in specimens from solid-organ transplant recipients for DNA by quantitative real-time polymerase chain reaction has been implemented in many diagnostic facilities. This technology provides rapid, accurate, and reproducible results for early detection, monitoring, and medical management of patients with these infections. Because these assays are becoming commonly used in clinical practice, the technical variables associated with specimen processing (e.g., nucleic acid extraction, gene target, and result reporting), amplification, and unique patient characteristics (e.g., age, sex, underlying diseases, immune status, and immunosuppressive regimens received) are factors that may influence the understanding and interpretation of test results. We emphasize the need for standardization of existing variables through parallel comparative and proficiency testing, uniform units for expressing results, to provide for clinical correlation with the results of these molecular assays.

Polyomavirus polymerase chain reaction as a surrogate marker of polyomavirus-associated nephropathy.

BACKGROUND: Polyomavirus-associated nephropathy (PVAN) is a significant cause of allograft loss after renal transplantation. A noninvasive assay that can guide the evaluation of PVAN would be of clinical value. We compared the utility of BK virus (BKV) polymerase chain reaction (PCR) and urine cytology in screening for concurrent PVAN. METHODS: We used PCR to test urine and plasma samples from renal recipients simultaneously for BKV DNA. Additionally, we tested urine samples for decoy cells. Sample results were correlated with biopsy-proven PVAN. Receiver-operator characteristic curves were used to determine viral load thresholds associated with concurrent PVAN. RESULTS: In this cross-sectional study, BKV viruria, viremia, and urinary decoy cells were detected in 24%, 9%, and 13% of renal recipients, respectively. Among 114 patients who had renal allograft biopsy, four (3.5%) were diagnosed with PVAN. Using pathology as gold standard for the diagnosis of PVAN, BKV viremia threshold of >1.6E+04 copies/mL had 100% sensitivity, 96% specificity, 50% positive predictive value, and 100% negative predictive value. A BKV viruria threshold of >2.5E+07 copies/mL had 100% sensitivity, 92% specificity, 31% positive predictive value, and 100% negative predictive value. In contrast, urine decoy cells had 25% sensitivity, 84% specificity, 5% positive predictive value, and 97% negative predictive value for the diagnosis of concurrent PVAN. CONCLUSION: BKV PCR may be a clinically useful noninvasive test to identify renal recipients with concurrent PVAN. BKV DNA >1.6E+04 copies/mL of plasma and >2.5E+07 copies/mL of urine were highly associated with concurrent PVAN whereas a negative PCR test makes the diagnosis of PVAN highly unlikely.

Duration of influenza A virus shedding in hospitalized patients and implications for infection control.

OBJECTIVE: To assess the duration of shedding of influenza A virus detected by polymerase chain reaction (PCR) and cell culture among patients hospitalized with influenza A virus infection. SETTING: Mayo Clinic (Rochester, Minnesota) hospitals that cater to both the community and referral populations. METHODS: Patients 18 years old and older who were hospitalized between December 1, 2004, and March 15, 2005, with a laboratory-confirmed (ie, PCR-based) diagnosis of influenza A virus infection were consecutively enrolled. Additional throat swab specimens were collected at 2, 3, 5, and 7 days after the initial specimen (if the patient was still hospitalized). All specimens were tested by PCR and culture (both conventional tube culture and shell vial assay). Information on demographic characteristics, date of symptom onset, comorbidities, immunosuppression, influenza vaccination status, and receipt of antiviral treatment was obtained by interview and medical record review. Patients were excluded if informed consent could not be obtained or if the date of symptom onset could not be ascertained. RESULTS: Of 149 patients hospitalized with influenza A virus infection, 50 patients were enrolled in the study. Most patients were older (median age, 76 years), and almost all (96%) had underlying chronic medical conditions. Of 41 patients included in the final analysis, influenza A virus was detected in 22 (54%) by PCR and in 12 (29%) by culture methods at or beyond 7 days after symptom onset. All 12 patients identified by culture also had PCR results positive for influenza A virus. CONCLUSION: Hospitalized patients with influenza A virus infection can shed detectable virus beyond the 5- to 7-day period traditionally considered the duration of infectivity. Additional research is needed to assess whether prolonging the duration of patient isolation is warranted to prevent nosocomial outbreaks during the influenza season.

Lack of association between Chlamydia pneumoniae seropositivity and aortic atherosclerotic plaques: a population-based transesophageal echocardiographic study.

OBJECTIVES: The objective of this study was to examine the relationship between Chlamydia pneumoniae seropositivity and aortic atherosclerotic plaques in the general population. BACKGROUND: Seroepidemiologic studies suggest that C pneumoniae infection plays a role in the pathogenesis of atherosclerosis. METHODS: Transesophageal echocardiography was performed in 385 subjects (median age 66 years, range 51 to 101 years; 53% men), a sample of the Olmsted County (Minnesota) population. The association between C pneumoniae immunoglobulin (Ig) G antibody titers and aortic atherosclerotic plaques was examined. RESULTS: Chlamydia pneumoniae IgG antibodies (titers >or=1:16) were detected in 287 subjects (74.5%): low titers (1:16 to 1:32) in 58 (15.1%), intermediate titers (1:64 to 1:128) in 144 (37.4%), and high titers (>or=1:256) in 85 subjects (22.1%). Antibody titers were not associated with the presence of aortic plaques after adjustment for age, gender, and smoking status (p = 0.64). Compared with titers <1:16, the adjusted odds ratios for aortic plaques were 1.46 (95% confidence interval [CI] 0.63 to 3.42) for low titers, 1.32 (95% CI 0.68 to 2.55) for intermediate titers, and 0.94 (95% CI 0.42 to 2.07) for high titers. Among the subgroup with plaques, antibody titers were not associated with the presence of plaques >or=4 mm thick (p = 0.99), plaques >or=6 mm (p = 0.49), or mobile debris (p = 0.71), after adjustment for age and smoking. CONCLUSIONS: Chlamydia pneumoniae IgG antibody titers are not associated with the presence or severity of aortic atherosclerosis in the general population. These observations do not support a role for C pneumoniae infection in the initiation or progression of atherosclerosis.

The pathogenesis of hepatitis C virus is influenced by cytomegalovirus.

We investigated the effect of beta-herpesviruses on allograft failure and mortality, hepatitis C virus (HCV) replication, and liver histologic characteristics among 92 HCV-infected liver transplant recipients. Reactivation of cytomegalovirus (CMV) but not of human herpesvirus 6 (HHV-6) was independently associated with allograft failure and mortality (risk ratio, 3.71; 95% confidence interval, 1.64-8.39); allograft failure and mortality was observed in 48% of patients with CMV disease, 35% of patients with subclinical CMV infection, and 17% of patients without CMV infection (P=.0275). CMV reactivation was highly predictive of mortality (P<.001), regardless of whether it remained subclinical or evolved into CMV disease. Patients with CMV disease had a higher fibrosis stage (P=.05) and had a trend toward a higher hepatitis activity index (P=.10) and HCV load (P=.10) at 16 weeks after liver transplantation. The pathogenesis of HCV is influenced by its interaction with CMV but not with HHV-6.

Inflammation, infection, and aortic valve sclerosis; Insights from the Olmsted County (Minnesota) population.

Atherosclerosis-related mechanisms, including inflammation and possibly infection, are likely to be involved in the pathogenesis of calcific aortic valve disease. The purpose of this study was to examine whether systemic inflammatory markers and Chlamydia pneumoniae seropositivity are associated with aortic valve sclerosis (AVS) in a sample of the general population. Transesophageal echocardiography was performed in 381 subjects (median age: 67 years, range: 51-101; 52% men), a sample of the adult population in Olmsted County, Minnesota. The associations between systemic inflammatory markers (blood counts, including white blood cells differential counts, fibrinogen, and high-sensitivity C-reactive protein [hs-CRP]), C. pneumoniae immunoglobulin G (IgG) antibody titers, and AVS were examined. AVS was present in 140 subjects (37% of the population). After adjustment for age, sex, and smoking status: (1). hs-CRP was associated with AVS (odds ratio: 1.20 per two-fold increase in hs-CRP; 95% confidence interval: 1.01-1.43; P = 0.04) but this association was not significant after adjustment for additional risk factors for AVS, including body mass index (P = 0.52). (2). Blood counts and fibrinogen were not associated with AVS (P-values >0.30). (3). C. pneumoniae IgG antibody titers (low [1:16-1:32], intermediate [1:64-1:128], or high [>or=1:256] titers, compared with titers <1:16) were not associated with AVS (P = 0.21). In conclusion, hs-CRP is weakly associated with AVS, an association that is not independent of other AVS risk factors. Blood counts, fibrinogen, and C. pneumoniae seropositivity are not associated with AVS. These findings suggest that other non-inflammatory non-infectious mechanisms are likely to have a role in the pathogenesis of calcific aortic valve disease.

The clinical use of various blood compartments for cytomegalovirus (CMV) DNA quantitation in transplant recipients with CMV disease.

BACKGROUND: The quantitation of cytomegalovirus (CMV) DNA is a cornerstone in the management of CMV disease in transplant recipients. However, a consensus as to what is the optimal blood compartment for the detection and quantitation of CMV DNA in peripheral blood is nonexistent. METHODS: With an automated quantitative assay, we have simultaneously quantified the CMV DNA load in whole blood (WB), plasma (PL), peripheral blood leukocytes (PBL), and peripheral blood mononuclear cells (PBMC) in 319 samples from 17 transplant recipients with 19 episodes of CMV disease that were treated with 2 weeks of intravenous ganciclovir. RESULTS: Higher levels of CMV DNA were observed in WB than PL (PL minus WB mean difference, 0.67 log; 95% confidence interval, -1.02 to -0.32; P=0.0009). This observation was most evident before treatment with intravenous ganciclovir (pretreatment geometric mean CMV DNA was 45,412 copies per ml of WB vs. 14,995 copies per ml of PL). In contrast, the CMV DNA levels between PBL and PBMC were highly comparable throughout the course of CMV disease and its treatment. Intravenous ganciclovir exerted a uniform effect on the four blood compartments with no statistically significant difference in the degree and rate of CMV DNA decline between WB and PL and between PBL and PBMC. CONCLUSIONS: Although our study demonstrates the adequacy of all blood compartments for CMV DNA quantification, the higher sensitivity of WB and its yield of higher CMV DNA render it an optimal sample for monitoring CMV DNA load during CMV disease in immunocompromised patients.

SARS: epidemiology, clinical presentation, management, and infection control measures.

Severe acute respiratory syndrome (SARS) is a recently recognized febrile respiratory illness that first appeared in southern China in November 2002, has since spread to several countries, and has resulted in more than 8000 cases and more than 750 deaths. The disease has been etiologically linked to a novel coronavirus that has been named the SARS-associated coronavirus. It appears to be spread primarily by large droplet transmission. There is no specific therapy, and management consists of supportive care. This article summarizes currently available information regarding the epidemiology, clinical features, etiologic agent, and modes of transmission of the disease, as well as infection control measures appropriate to contain SARS.

Avian influenza: a new pandemic threat?

In December 2003, the largest outbreak of highly pathogenic avian influenza H5N1 occurred among poultry in 8 Asian countries. A limited number of human H5N1 infections have been reported from Vietnam and Thailand, with a mortality rate approaching 70%. Deaths have occurred in otherwise healthy young individuals, which is reminiscent of the 1918 Spanish influenza pandemic. The main presenting features were fever, pneumonitis, lymphopenia, and diarrhea. Notably, sore throat, conjunctivitis, and coryza were absent. The H5N1 strains are resistant to amantadine and rimantadine but are susceptible to neuraminidase inhibitors, which can be used for treatment and prophylaxis. The widespread epidemic of avian influenza in domestic birds increases the likelihood for mutational events and genetic reassortment. The threat of a future pandemic from avian influenza is real. Adequate surveillance, development of vaccines, outbreak preparedness, and pandemic influenza planning are important. This article summarizes the current knowledge on avian influenza, including the virology, epidemiology, diagnosis, and management of this emerging disease.

Confronting bioterrorism: physicians on the front line.

The events surrounding September 11, 2001, and its aftermath have compelled the public health and medical community to face the hitherto unfamiliar reality of bioterrorism. Physicians and public health personnel are frontline soldiers in this new form of warfare. This article provides a general overview of the pathophysiology, clinical presentation, diagnosis, and management of patients infected with the 6 highest priority agents that could potentially be used in bioterrorism. The diseases discussed include anthrax, smallpox, tularemia, plague, botulism, and viral hemorrhagic fevers. Despite the unpredictable nature of bioterrorism, disaster preparedness and knowledge of essential diagnostic and epidemiological principles can contribute substantially toward combating this new threat.

Detection of vaccinia virus, herpes simplex virus, varicella-zoster virus, and Bacillus anthracis DNA by LightCycler polymerase chain reaction after autoclaving: implications for biosafety of bioterrorism agents.

OBJECTIVE: To determine whether autoclaving suspensions of vaccinia virus, herpes simplex virus (HSV), varicella-zoster virus (VZV), and Bacillus anthracis inactivate infectivity of these agents but allow detection of target DNA by LightCycler polymerase chain reaction (PCR). MATERIAL AND METHODS: Swabs were inserted into tubes containing serial 10-fold dilutions (10(-1) to 10(-5); 500 microL; 6 samples per dilution) of vaccinia virus, HSV, VZV, or a single suspension of 10(8) colony-forming units of B anthracis (2 samples). One half of the samples were autoclaved, and the remainder were not. An aliquot of each not autoclaved sample served as a control for infectivity. RESULTS: Autoclaving swabs saturated with suspensions of vaccinia virus, HSV, or VZV eliminated the infectivity of these agents; however, DNA was detectable in most autoclaved samples in dilutions of 10(-1) to 10(-4) by LightCycler PCR. All not autoclaved specimens were detected by culture (infectivity) except for VZV and, in most dilutions of 10(-1) to 10(-3), by assay of target DNA by LightCycler PCR. Similarly positive results were obtained for PCR assessment of sporulated B anthracis. CONCLUSIONS: Standard autoclaving procedures eliminated the infectivity of viruses (and B anthracis), but target DNA was often retained for detection by LightCycler PCR. Current recommendations indicate that the laboratory diagnosis of smallpox virus infection be performed only within Biosafety Level 4 facilities. We suggest that, in addition to the requirement for immediate coordination with public health officials, the federal government consider expanding the existing guidelines for processing these specimens to encourage immediate collection, autoclaving, and testing by LightCycler PCR to differentiate smallpox virus from other dermal pathogens such as HSV and VZV by specific qualified laboratories.

Application of rapid-cycle real-time polymerase chain reaction for the detection of microbial pathogens: the Mayo-Roche Rapid Anthrax Test.

Rapid-cycle real-time polymerase chain reaction has immediate and important implications for diagnostic testing in the clinical microbiology laboratory. In our experience this novel testing method has outstanding performance characteristics. The sensitivities for detecting microorganisms frequently exceed standard culture-based assays, and the time required to complete the assays is considerably shorter than that required for culture-based assays. We describe the principle of real-time polymerase chain reaction and present clinical applications, including the detection of Bacillus anthracis, the causative agent of anthrax. This latter test is commercially available as the result of a collaborative venture between Mayo Clinic and Roche Applied Science, hence the designation The Mayo-Roche Rapid Anthrax Test.

Comparison of the VERSANT HCV RNA qualitative assay (transcription-mediated amplification) and the COBAS AMPLICOR hepatitis C virus test, version 2.0, in patients undergoing interferon-ribavirin therapy.

Hepatitis C virus (HCV)-infected patients were tested for the presence of HCV RNA using two qualitative assays at various time points during interferon-ribavirin therapy. Among patients treated for 48 weeks, transcription-mediated amplification and the COBAS AMPLICOR Hepatitis C Virus Test results at Week 24 predicted subsequent virologic non-response or virologic relapse in 12/15 (80%) and 8/15 (53%) patients, respectively.

Comparison of line probe assay and DNA sequencing of 5' untranslated region for genotyping hepatitis C virus: description of novel line probe patterns.

We compared a commercial line probe assay (INNO-LiPA HCV II, Innogenetics, N.V., Ghent, Belgium, distributed by Bayer Diagnostics) to an in-house 5' untranslated region direct DNA sequencing method for genotyping hepatitis C virus (HCV). Initial evaluation demonstrated that the INNO-LiPA HCV II assay and sequencing assay assigned the same genotype for 110/132 (83.3%) patient specimens (98 subtype and 12 genotype only identifications). Following the initial evaluation, the INNO-LiPA HCV II assay was used routinely to genotype HCV from patient specimens submitted to our laboratory for genotyping (n = 1,739). During this second part of the study, novel line probe patterns have been noted and interpreted using the in-house direct sequencing assay. Reactivity at bands 1, 2, 3, 4, 5 and 8 (n = 4) or 1, 2, 3, 4, 6 and 7 (n = 2) represented HCV genotype 1. Reactivity at bands 1, 2, 5 and 9 (n = 1) represented HCV genotype 2. Reactivity at bands 1, 2, 5, 9 and 16 (n = 1) represented HCV genotype 4. Reactivity at bands 1, 2, 5, 9, 10, 11 (weak band) and 12 (n = 118) most likely represented HCV genotype 2b. This information should be of use to INNO-LiPA HCV II assay users.

Viral detection using a multiplex polymerase chain reaction-based assay in outpatients with upper respiratory infection.

We evaluated a commercial multiplex polymerase chain reaction (PCR) assay in a cross-sectional study among 81 adult and pediatric outpatients-40 cases with upper respiratory infection symptoms and 41 asymptomatic controls-from February to April 2008. Two specimens (throat swab and nasal swab) from each participant were tested using the EraGen MultiCode-PLx Respiratory Virus Panel that detects 17 viral targets. Throat swabs were also tested for Group A Streptococcus (GAS) by PCR. Respiratory viruses were detected in 22/40 (55%) cases and in 3/41 (7%) controls (P < 0.001). GAS was detected in 10 (25%) cases; GAS and respiratory virus co-infection was found in 4 (10%). Agreement between nasal and throat swabs for viral detection was 69% in cases and 95% in controls. Of 22 cases with a detectable virus, 12 (54%) were picked up by only 1 (throat or nasal) specimen, and the detection rate was increased by combining results of nasal and throat swab testing.