Impact of Pool Testing in Detection of Asymptomatic Patients with COVID-19.

OBJECTIVE: During the current pandemic, COVID-19 has been detected in patients using real-time reverse transcriptase-polymerase chain reaction (RT-PCR) that confirms the presence of SARS-CoV-2 RNA. The demand for increased testing, particularly for asymptomatic individuals required alternative approaches to single-patient RT-PCR testing, such as pooling. METHODS: This study explored the impact of dilution on the detectability of SARS-CoV-2 in asymptomatic patients using RT-PCR and demonstrated that pooling can be effective in low prevalence populations. RESULTS: The RT-PCR results for the 3:1, 5:1, and 7:1 aliquot samples showed little differences in CT values, confirming detection capability at these dilutions. CONCLUSION: Based on the results of the present study, a pooled approach with up to 5:1 sample aliquots and using the current RT-PCR methodology likely will detect SARS CoV2 RNA among asymptomatic patients.

Role of HrpA in biofilm formation of Neisseria meningitidis and regulation of the hrpBAS transcripts.

Two-partner secretion systems of gram-negative organisms are utilized in adherence, invasion, and biofilm formation. The HrpAB proteins of Neisseria meningitidis are members of a two-partner secretion system, and HrpA is established as being important to adherence and intracellular escape. This study set out to determine the expression pattern of members of the hrpBAS putative operon and to find a functional role for the HrpA protein. The upregulation of these genes was found in situations of anaerobiosis and cell contact. These observations prompted the study of the function of HrpA in biofilms on human bronchial epithelial cells. HrpA mutants in encapsulated and unencapsulated NMB strains demonstrated biofilm growth equivalent to that of the wild-type strain at 6 h but a decreased ability to form biofilms at 48 h. Biofilms formed by hrpA mutants for 48 h on collagen-coated coverslips demonstrated significant reductions compared to those of wild-type strains. Taken together, these observations imply a role for HrpA in the biofilm structure. Further analysis demonstrated the presence of HrpA on the surface of the bacterium.

Using fee-for-service testing to generate revenue for the 21st century public health laboratory.

OBJECTIVES: The decrease in appropriations for state public health laboratories (SPHLs) has become a major concern as tax revenues and, subsequently, state and federal funding, have decreased. These reductions have forced SPHLs to pursue revenue-generating opportunities to support their work. We describe the current state of funding in a sampling of SPHLs and the challenges these laboratories face as they implement or expand fee-for-service testing. METHODS: We conducted surveys of SPHLs to collect data concerning laboratory funding sources, test menus, fee-for-service testing, and challenges to implementing fee-for-service testing. RESULTS: Most SPHLS receive funding through three revenue sources: state appropriation, federal funding, and fee-for-service testing (cash funds). Among SPHLs, state appropriations ranged from $0 to more than $6 per capita, federal funding ranged from $0.10 to $5 per capita, and revenue from fee-for-service testing ranged from $0 to $4 per capita. The tests commonly performed on a fee-for-service basis included assays for sexually transmitted diseases, mycobacterial cultures, newborn screening, and water testing. We found that restrictive legislation, staffing shortages, inadequate software for billing fee-for-service testing, and regulations on how SPHLs use their generated revenue are impediments to implementing fee-for-service testing. CONCLUSIONS: Some SPHLs are considering implementing or expanding fee-for-service testing as a way to recapture funds lost as a result of state and federal budget cuts. This analysis revealed many of the obstacles to implementing fee-for-service testing in SPHLs and the potential impact on SPHLs of continued decreases in funding.

The frequency and seasonality of influenza and other respiratory viruses in Tennessee: two influenza seasons of surveillance data, 2010-2012.

BACKGROUND: In 2010, the Tennessee Department of Health, in collaboration with the Centers for Disease Control and Prevention (CDC), expanded influenza surveillance in Tennessee to include other respiratory viruses. OBJECTIVES: To determine the prevalence and seasonality of influenza and other respiratory viruses during the influenza seasons of 2010-2012. METHODS: Nasal and nasopharangeal swabs/washings from persons with influenza-like illness were collected across Tennessee. Influenza and other respiratory viruses were identified using a molecular-based respiratory virus panel. Influenza A positives were subtyped using real-time PCR according to the CDC protocol. Data were analyzed to describe frequency and seasonality of circulating strains. RESULTS: Of the 933 positive specimens, 60·3% were identified as influenza viruses, 19·8% rhinovirus/enterovirus, 8·6% respiratory syncytial virus (RSV), 5·8% metapneumovirus, 3·0% adenovirus, and 2·5% parainfluenza viruses. In the 2010-2011 season, influenza B was prominent during weeks 48-3, while influenza A(H1N1) was most frequently identified during weeks 4-10. Influenza A(H3N2) was present at lower levels during weeks 48-17. However, in the 2011-2012 season, overall numbers of influenza cases were reduced and influenza A (H3N2) was the most abundant influenza strain. The expanded surveillance for other respiratory viruses noted an increase in identified specimens from the first to the second season for adenovirus, metapneumovirus, RSV, and rhinovirus/enterovirus. CONCLUSIONS: This study provides data of the influenza strains in circulation in Tennessee. It also establishes a baseline and time of year to expect other respiratory viruses that will aid in detecting outbreaks of non-influenza respiratory viruses in Tennessee.

Methods for studying Neisseria meningitidis biofilms.

Neisseria meningitidisis an organism whose environmental niche is limited to the human host. It can frequently colonize the human nasopharynx and has the ability to cause severe systemic infections. These infections can be sporadic, endemic or occur in outbreaks associated with more virulent meningococcal strains. Studies have demonstrated that the meningococcus can form biofilms both in vivo and ex vivo. In this chapter, we discuss methods to establish biofilms in the laboratory for in-depth biochemical, genetic, or microscopic studies.

Clinical and laboratory evidence for Neisseria meningitidis biofilms.

Neisseria meningitidis is the etiologic agent of meningococcal meningitis. Carriage of the organism is approximately 10% while active disease occurs at a rate of 1:100,000. Recent publications demonstrate that N. meningitidis has the ability to form biofilms on glass, plastic or cultured human bronchial epithelial cells. Microcolony-like structures are also observed in histological sections from patients with active meningococcal disease. This review investigates the possible role of meningococcal biofilms in carriage and active disease, based on the laboratory and clinical aspects of the disease.

Biofilm formation on human airway epithelia by encapsulated Neisseria meningitidis serogroup B.

Neisseria meningitidis is the etiologic agent of meningococcal meningitis. We compared 48-h biofilm formation by N. meningitidis serogroup B strains NMB, MC58, C311 and isogenic mutants defective in capsule formation on SV-40 transformed human bronchial epithelial (HBE) cells in a flow cell. We demonstrated that strains NMB and NMB siaA-D were defective in biofilm formation over glass, and there was a partial rescue of biofilm growth for strain NMB on collagen-coated coverslips at 48 h. We demonstrated all three serogroup B strains form biofilms of statistically equivalent average height on HBE cells as their isogenic capsular mutants. Strain NMB also formed a biofilm of statistically equivalent biomass as the NMB siaA-D mutant on HBE cells at 6 and 48 h. These biofilms are significantly larger than biofilms formed over glass or collagen. Verification that strain NMB expressed capsule in biofilms on HBE cells was demonstrated by staining with 2.2.B, a monoclonal antibody with specificity for the serogroup B capsule. ELISA analysis demonstrated that strains MC58 and C311 also produced capsules during biofilm growth. These findings suggest that encapsulated meningococci can form biofilms on epithelial cells suggesting that biofilm formation may play a role in nasopharyngeal colonization.

Myxobacterial biodiversity in an established oak-hickory forest and a savanna restoration site.

The purpose of this study was to establish the myxobacterial biodiversity of an established oak-hickory forest and a savanna restoration site that has been cut and subsequently burned on four occasions between 1993 and 1998 in an attempt to restore the land to the native savanna ecosystem. Soil and bark samples were processed through standard methods specifically for myxobacteria and numbers and types of species were recorded for both locations. Species were identified through morphology of fruiting bodies, SDS-PAGE of whole cell protein profiles, and 16S rRNA gene sequences. Statistical analyses were employed and suggested that significantly greater numbers and types of myxobacteria are present on the bark of the trees in the established oak-hickory forest than the bark of trees in the savanna restoration site, while little difference in numbers and types of species were observed between the soil samples of the two locations.