Effective laboratory methods for identifying avian influenza virus (AIV) in wild bird populations are crucial to understanding the ecology of this pathogen. The standard method has been AIV isolation in chorioallantoic sac (CAS) of specific-pathogen-free embryonating chicken eggs (ECE), but in one study, combined use of yolk-sac (YS) and chorioallantoic membrane inoculation routes increased the number of virus isolations. In addition, cell culture for AIV isolation has been used. Most recently, real-time reverse transcriptase (RRT)-PCR has been used to detect AIV genome in surveillance samples. The purpose of this study was to develop a diagnostic decision tree that would increase AIV isolations from wild bird surveillance samples when using combinations of detection and isolation methods under our laboratory conditions. Attempts to identify AIV for 50 wild bird surveillance samples were accomplished via isolation in ECE using CAS and YS routes of inoculation, and in Madin-Darby canine kidney (MDCK) cells, and by AIV matrix gene detection using RRT-PCR. AIV was isolated from 36% of samples by CAS inoculation and 46% samples by YS inoculation using ECE, isolated from 20% of samples in MDCK cells, and detected in 54% of the samples by RRT-PCR. The AIV was isolated in ECE in 13 samples by both inoculation routes, five additional samples by allantoic, and 10 additional samples by yolk-sac inoculation, increasing the positive isolation of AIV in ECE to 56%. Allantoic inoculation and RRT-PCR detected AIV in 14 samples, with four additional samples by allantoic route alone and 13 additional samples by RRT-PCR. Our data indicate that addition of YS inoculation of ECE will increase isolation of AIV from wild bird surveillance samples. If we exclude the confirmation RT-PCR test, cost analysis for our laboratory indicates that RRT-PCR is an economical choice for screening samples before doing virus isolation in ECE if the AIV frequency is low in the samples. In contrast, isolation in ECE via CAS and YS inoculation routes without prescreening by RRT-PCR was most efficient and cost-effective if the samples had an expected high frequency of AIV.

Purpose Despite multiple studies regarding modes of eyebrow measurement and movement over time, the lateral aspect of the brow has been relatively ignored in literature. Therefore, we arranged a study of the most lateral aspect of the eyebrow; in doing so, we hoped to ascertain the most practical line or angle of measurement. Methods In this cross-sectional study, adults aged 18 and older with no history of congenital or acquired periorbital or orbital pathology or surgery; brow tattooing or heavy plucking; phthisis; or strabismus were measured using a combination of in-office metrics and computer analysis. Subjects were asked to identify their ethnicity and country of origin. Models of age, gender, age and ethnicity were created. Results 1024 subjects were included (1944 eyes). Measurements of nasal ala to lateral brow (NALB), lateral brow plumb line (LBPL, the vertical line between the tail of the brow and a horizontal line extending from the lateral canthus), and angle from the mid-brow to the lateral brow tail all showed statistically significant decline over time. The angle and LBPL varied most by ethnicity. The angle narrowed approximately 3 degrees per 20 years, while the LBPL fell approximately 2.5 mm per 20 years. The NALB varied most by age, and fell approximately 3 mm per 20 years. Conclusions The lateral tail of the brow descends with age. Measurements of its location and rate of change vary between genders and within ethnic groups. Two easily measured values – NALB and LBPL – can be used for pre-operative planning and post-operative documentation.