Microbial diversity and nitrogen-metabolizing gene abundance in backyard food waste composting systems.

AIMS: The microbial diversity of backyard compost piles is poorly understood compared to large-scale, highly regulated composting systems. The purpose of this study was the identification of the microbial community composition and associated change over time among three different backyard composting styles. METHODS AND RESULTS: Food waste was composted in a household backyard compost bin, a small-scale aerated windrow or a semi-aerated static pile. Samples were obtained from each sequential phase of the composting process for 16s rRNA sequencing and relationships between temperature, moisture and microbial communities were examined. The Bacilli dominated in the early phases of composting then transitioned to Proteobacteria in the later stages. Different bacterial species increased and decreased dramatically in different composting systems and at different phases of the composting process. We performed qPCR to quantify gene abundance of nirS to profile the nitrogen-metabolizing bacteria present in each composting system. Gene abundance of nirS varied with temperature, but peaked during the cooling phase in the aerated windrow. CONCLUSIONS: Although the phases of decomposition were not as distinct as large-scale regulated piles, the microbial diversity mirrored the appropriate phases. Interestingly, different backyard composting styles were marked by the predominance of certain bacterial species. In particular, nitrogen-metabolizing bacterial communities peaked in the later stages of decomposition. SIGNIFICANCE AND IMPACT OF THE STUDY: A profile of the compost microbiome yields important clues about how differences in backyard food waste composting systems influence bacterial species that may facilitate or hinder nitrogen metabolism.

SARS-CoV-2 antibody prevalence by industry, workplace characteristics, and workplace infection prevention and control measures, North Carolina, 2021 to 2022.

Background: The COVID-19 pandemic has disproportionately affected workers in certain industries and occupations, and the workplace can be a high risk setting for SARS-CoV-2 transmission. In this study, we measured SARS-CoV-2 antibody prevalence and identified work-related risk factors in a population primarily working at industrial livestock operations. Methods: We used a multiplex salivary SARS-CoV-2 IgG antibody assay to determine infection-induced antibody prevalence among 236 adult (≥18 years) North Carolina residents between February 2021 and August 2022. We used the National Institute for Occupational Safety and Health Industry and Occupation Computerized Coding System (NIOCCS) to classify employed participants' industry and compared infection-induced IgG prevalence by participant industry and with the North Carolina general population. We also combined antibody results with reported SARS-CoV-2 molecular test positivity and vaccination history to identify evidence of prior infection. We used logistic regression to estimate odds ratios of prior infection by potential work-related risk factors, adjusting for industry and date. Results: Most participants (55%) were infection-induced IgG positive, including 71% of animal slaughtering and processing industry workers, which is 1.5 to 4.3 times higher compared to the North Carolina general population, as well as higher than molecularly-confirmed cases and the only other serology study we identified of animal slaughtering and processing workers. Considering questionnaire results in addition to antibodies, the proportion of participants with evidence of prior infection increased slightly, to 61%, including 75% of animal slaughtering and processing workers. Participants with more than 1000 compared to 10 or fewer coworkers at their jobsite had higher odds of prior infection (adjusted odds ratio [aOR] 4.5, 95% confidence interval [CI] 1.0 to 21.0). Conclusions: This study contributes evidence of the severe and disproportionate impacts of COVID-19 on animal processing and essential workers and workers in large congregate settings. We also demonstrate the utility of combining non-invasive biomarker and questionnaire data for the study of workplace exposures.

Multidrug-resistant tuberculosis in children: A practical update on epidemiology, diagnosis, treatment and prevention.

Pediatric multidrug-resistant tuberculosis (MDR-TB) remains a significant global problem, and there are numerous barriers preventing children with MDR-TB from being identified, confirmed with microbiologic tests, and treated with a safe, practical, and effective regimen. However, several recent advances in diagnostics and treatment regimens have the promise to improve outcomes for children with MDR-TB. We introduce this review with two cases that exemplify both the challenges in management of MDR-TB in children, but also the potential to achieve a positive outcome. More than 30,000 cases of MDR-TB per year are believed to occur in children but less than 5% are confirmed microbiologically, contributing to poorer outcomes and excess mortality. Rapid molecular-based testing that provides information on rifampin susceptibility is increasingly globally available and recommended for all children suspected of TB disease--but remains limited by challenges obtaining appropriate samples and the paucibacillary nature of most pediatric TB. More complex assays allowing better characterization of drug-resistant isolates are emerging. For children diagnosed with MDR-TB, treatment regimens have traditionally been long and utilize multiple drugs associated with significant side effects, particularly injectable agents. Several new or repurposed drugs including bedaquiline, delamanid, clofazimine and linezolid now allow most treatment regimens to be shorter and all-oral. Yet data to support short, all-oral, novel regimens for young children containing pretomanid remain insufficient at present, and there is a compelling need to conduct pediatric trials of promising therapeutics and MDR-TB treatment regimens.

SARS-CoV-2 Antibody Prevalence among Industrial Livestock Operation Workers and Nearby Community Residents, North Carolina, 2021 to 2022.

Industrial livestock operations (ILOs), particularly processing facilities, emerged as centers of coronavirus disease 2019 (COVID-19) outbreaks in spring 2020. Confirmed cases of COVID-19 underestimate true prevalence. To investigate the prevalence of antibodies against SARS-CoV-2, we enrolled 279 participants in North Carolina from February 2021 to July 2022: 90 from households with at least one ILO worker (ILO), 97 from high-ILO intensity areas (ILO neighbors [ILON]), and 92 from metropolitan areas (metro). More metro (55.4%) compared to ILO (51.6%) and ILON participants (48.4%) completed the COVID-19 primary vaccination series; the median completion date was more than 4 months later for ILO compared to ILON and metro participants, although neither difference was statistically significant. Participants provided a saliva swab we analyzed for SARS-CoV-2 IgG using a multiplex immunoassay. The prevalence of infection-induced IgG (positive for nucleocapsid and receptor binding domain) was higher among ILO (63%) than ILON (42.9%) and metro (48.7%) participants (prevalence ratio [PR], 1.38; 95% confidence interval [CI], 1.06 to 1.80; reference category ILON and metro combined). The prevalence of infection-induced IgG was also higher among ILO participants than among an Atlanta health care worker cohort (PR, 2.45; 95% CI, 1.80 to 3.33) and a general population cohort in North Carolina (PRs, 6.37 to 10.67). The infection-induced IgG prevalence increased over the study period. Participants reporting not masking in public in the past 2 weeks had higher infection-induced IgG prevalence (78.6%) than participants reporting masking (49.3%) (PR, 1.59; 95% CI, 1.19 to 2.13). Lower education, more people per bedroom, Hispanic/Latino ethnicity, and more contact with people outside the home were also associated with higher infection-induced IgG prevalence. IMPORTANCE Few studies have measured COVID-19 seroprevalence in North Carolina, especially among rural, Black, and Hispanic/Latino communities that have been heavily affected. Antibody results show high rates of COVID-19 among industrial livestock operation workers and their household members. Antibody results add to evidence of health disparities related to COVID-19 by socioeconomic status and ethnicity. Associations between masking and physical distancing with antibody results also add to evidence of the effectiveness of these prevention strategies. Delays in the timing of receipt of COVID-19 vaccination reinforce the importance of dismantling vaccination barriers, especially for industrial livestock operation workers and their household members.

Early synthesis of specific cytoplasm proteins is correlated with the rate of exit of lymphocytes from the resting state.

We investigated the initiation of synthesis of proteins in human lymphocytes exposed to the mitogen phytohemagglutinin (PHA) for 6 h. Radiolabeled proteins in three subcellular fractions, cytoplasmic, nuclear salt wash, and nuclear, were separated on polyacrylamide gels. Compared with cells incubated for the same time in the absence of PHA only two cytoplasmic proteins of Mr 51 and 60 kd showed increased synthesis in a dose-dependent fashion. Synthesis of the 60-kd protein shows the strongest correlation with rate of entry into the first S phase and with rate of cellular aggregation. Thus, the 60-kd protein appears to be a major early response-associated protein for entry of lymphocytes into the first S phase after PHA stimulation.

Evidence for a novel signal transduction pathway activated by platelet-derived growth factor and by double-stranded RNA.

Platelet-derived growth factor (PDGF) and the synthetic double-stranded RNA poly(I).poly(C) [poly(I.C)] stimulate transcription of the JE gene in BALB/c-3T3 fibroblasts. The response of JE to poly(I.C) does not appear to be channeled through any known component of the PDGF receptor signal transduction apparatus. In addition, JE sequences upstream of the transcription start site are devoid of previously identified poly(I.C)-responsive elements, such as those found in the beta-interferon gene. These data suggest that a novel signal transduction pathway regulates the JE response to PDGF and double-stranded RNA. The c-myc and c-fos proto-oncogenes also respond to this pathway but with poor efficiency. However, this pathway operates very efficiently on other PDGF-inducible genes that encode the secretory proteins KC and M-CSF.

Drivers of professional mobility in the Northern Territory: dental professionals.

INTRODUCTION: Attracting and retaining an efficient allied health workforce is a challenge faced by communities in Australia and overseas. High rates of staff turnover in the professional workforce diverts resources away from core business and results in the loss of valuable skills and knowledge. Understanding what attracts professionals to a particular place, and why they leave, is important for developing effective strategies to manage turnover and maximise workforce productivity. The Northern Territory (NT) faces particular workforce challenges, in part because of its geographic location and unusual demography. Do these factors require the development of a tailored approach to recruitment and retention? This article reports on a study undertaken to examine the motivations for coming to, staying in and leaving the NT for dental professionals, and the implications of results on workforce management practices. METHODS: In 2006, dentists, dental specialists, dental therapists and dental hygienists who were working or had worked in the NT, Australia, in the recent past were surveyed to collect demographic and workforce data and to establish the relative importance of social and work-related factors influencing their migration decisions. Multivariate logistic regression models were generated to describe the demographic characteristics of dental professionals who stayed in the NT for more than 5 years and to analyse why dental professionals left. The analyses, based on a 42% response rate, explained 60-80% of the variation in responses. RESULTS: Generally dental professionals who had stayed for more than 5 years were older, had invested in the purchase of homes and were more involved in social and cultural activities. Those who moved to the NT as a result of financial incentives or who had strong expectations that working in the NT would be an exciting, novel experience tended to stay for no more than 5 years, often leaving because they found the work environment too stressful. In contrast, those who stayed longer came because they had existing social networks and were familiar with the NT environment, staying primarily because they have enjoyed the NT lifestyle, particularly the sense of community and the opportunities available through living in smaller centres. CONCLUSION: There are benefits in actively engaging newly recruited professionals and their families in social networks. Work related stress and departure was associated with administrative deficiencies within the management system. Despite the NT's unusual demographic profile, the factors influencing recruitment and retention are not markedly different from those reported elsewhere.

Expression of a deletion mutant of the E2F1 transcription factor in fibroblasts lengthens S phase and increases sensitivity to S phase-specific toxins.

To better understand how the E2F1 transcription factor contributes to the process of cell proliferation, NIH-3T3 cell lines were generated that constitutively express either the wild-type E2F1 protein or an amino terminal deletion mutant, termed E2F1d87. Proliferating E2F1d87-expressing cells exhibit a significant lengthening of S phase relative to control and E2F1 cell lines and are hypersensitive to the cytotoxic effects of the S phase-specific antitumor drug camptothecin. This sensitivity is associated with an increase in drug-induced p53 and WAF1 levels. The E2F1 and E2F1d87 cell lines are both able to initiate, but not complete, S phase under conditions of serum starvation. However, quantitation of DNA synthesis, during culture in serum-deprived media, indicates that the E2F1d87 cell line synthesizes more DNA/cell as compared to the E2F1 cell line. Consistent with this relative increase in DNA synthesis, the E2F1d87 cell line undergoes camptothecin-induced apoptosis when cultured under conditions of serum starvation, while the control and E2F1 cell lines are unaffected by drug treatment under the same conditions. Thus, the sensitivity of the E2F1d87 cell line to camptothecin is not dependent on cell proliferation. The data presented here suggest that cell cycle parameters can be manipulated in order to enhance sensitivity of a cell to the toxic effects of specific chemotherapeutic agents.

Regulation of c-myc transcription in vitro: dependence on the guanine-rich promoter element ME1a1.

P2 is the major promoter for the murine c-myc proto-oncogene. The cis-acting elements that are required for initiation of transcription from P2 have not been well defined. In this report elements involved in initiating transcription from P2 were analysed in an in vitro system. In addition to a consensus TATA element at position -28, a guanine-rich element exists at position -48. This element, termed ME1a1, increases transcription initiation when inserted into a deletion mutant that lacks it. When mutations are engineered into ME1a1 it no longer acts to increase the level of initiation. Gel-shift and DNAase I footprinting analysis indicate that Me1a1 binds a protein. ME1a1 does not show any striking similarity to other promoter elements and therefore may be a novel cis-acting element.

Detailed analysis of the basic domain of the E2F1 transcription factor indicates that it is unique among bHLH proteins.

The E2F1 transcription factor binds to sites within the promoters of a number of cell cycle regulated genes through a basic-helix-loop-helix motif (bHLH). It is shown here that the basic region of E2F1 is distinct from that of all other bHLH proteins. The center of the basic region contains a helix breaking proline-glycine pair, (P122, G123), implying a turn within this region. This is in contrast to the known bHLH containing proteins where the basic region is alpha-helical. Substitution of P122 and G123 with alanines results in a significant reduction in DNA binding levels, with the predicted formation of an alpha-helix. Also in contrast to other bHLH proteins, mutations generated in conserved basic residues of E2F1 do not effect DNA binding. In addition, a single leucine (191) between helix no. 2 and the leucine zipper is required for DNA binding while the leucine zipper itself is not necessary. Finally, E2F1 interacts with all of the G-residues in the sequence GGCGGGAAA while the A-residues are not required for DNA binding. The uniqueness of the E2F1 DNA binding domain is likely to play a role in its binding a DNA site that is distinct from that of all other bHLH proteins (CACGTG).

Three distinct elements within the murine c-myc promoter are required for transcription.

We have undertaken a detailed analysis of the cis-acting elements that are required for optimal transcription initiation from P2, the major promoter of the murine c-myc gene. We find that three elements contribute to promoter strength, termed ME1a2, E2F and ME1a1 at positions -85, -64 and -46 respectively (relative to P2). Individually the elements are weak, but combined they contribute to full promoter activity, all acting in a positive fashion. The E2F element is the site at which the SV40 large T antigen transactivates the c-myc promoter. However, transactivation requires the presence of the ME1a2 or ME1a1 elements in addition to E2F. By a number of criteria it appears that the ME1a2 and ME1a1 elements bind the same or a very closely related protein (monomer Mr 94,000). It was found that Hela cells contain a novel factor that is capable of binding to the ME1a1 and ME1a2 elements but only in the presence of the protein-dissociating agents deoxycholate or formamide. Finally, the E2F factor binds DNA both as a monomer and as a multiprotein complex; the latter is cell type specific. Both types of bound E2F factor form less stable protein-DNA complexes than the ME1a2/ME1a1 factor.

A three-dimensional nanofibrous scaffold for cartilage tissue engineering using human mesenchymal stem cells.

The utilization of adult stem cells in tissue engineering is a promising solution to the problem of tissue or organ shortage. Adult bone marrow derived mesenchymal stem cells (MSCs) are undifferentiated, multipotential cells which are capable of giving rise to chondrocytes when maintained in a three-dimensional culture and treated with members of the transforming growth factor-beta (TGF-beta) family of growth factors. In this study, we fabricated a nanofibrous scaffold (NFS) made of a synthetic biodegradable polymer, poly(-caprolactone) (PCL), and examined its ability to support in vitro chondrogenesis of MSCs. The electrospun PCL porous scaffold was constructed of uniform, randomly oriented nanofibers with a diameter of 700 nm, and structural integrity of this scaffold was maintained over a 21-day culture period. MSCs cultured in NFSs in the presence of TGF-beta1 differentiated to a chondrocytic phenotype, as evidenced by chondrocyte-specific gene expression and synthesis of cartilage-associated extracellular matrix (ECM) proteins. The level of chondrogenesis observed in MSCs seeded within NFSs was comparable to that observed for MSCs maintained as cell aggregates or pellets, a widely used culture protocol for studying chondrogenesis of MSCs in vitro. Due to the physical nature and improved mechanical properties of NFSs, particularly in comparison to cell pellets, the findings reported here suggest that the PCL NFS is a practical carrier for MSC transplantation, and represents a candidate scaffold for cell-based tissue engineering approaches to cartilage repair.

Positive regulation of human alpha 1 (I) collagen promoter activity by transcription factor Sp1.

Analysis of the regulatory promoter region of the human alpha 1 (I) collagen-encoding gene (COL1A1) gene indicated the presence of G+C-rich sequence elements that are potential binding sites for the transcription factor Sp1. As a step toward understanding transcriptional regulation of the human COL1A1, we examined Sp1 binding in the promoter region using DNase I footprinting, and analyzed the effect of Sp1 expression on COL1A1 promoter activity in transiently transfected Drosophila melanogaster cells in vivo. The results indicated that recombinant human Sp1 interacted specifically with two G+C-rich sequences within the COL1A1 promoter. Binding of factors in nuclear extracts prepared from human dermal fibroblasts to a 22-nucleotide deoxyribonucleotide (oligo) spanning the 5' G+C-rich sequence required Zn2+, and was abolished by excess Sp1 consensus binding site oligos, or by anti-Sp1 antibodies. Studies in which a series of progressively 5'-deleted COL1A1 promoter::cat constructs were co-expressed with an Sp1 expression plasmid in a cellular background devoid of Sp1 homology demonstrated that Sp1 markedly enhanced the COL1A1 promoter activity. These results suggest that the transcriptional activity of the human COL1A1 can be positively regulated by Sp1.

Tumor necrosis factor and gamma-interferon repress transcription from the c-myc P2 promoter by reducing E2F binding activity.

Transcription of the c-myc gene in HeLa cells has been shown to be repressed by the combined action of tumor necrosis factor (TNF) and gamma-interferon (gamma-INF). Shown here, these two cytokines inhibit proliferation of Hela cells with a coordinate inhibition of c-myc gene expression. It was found that these two cytokines exert their effects on the more proximal region of the c-myc P2 promoter. Using c-myc promoter:CAT constructs, it was found that the combined action of these cytokines significantly repress transcription from P2. This repression occurred through the E2F site within the promoter and not the ME1a2 or ME1a1 sites. However, these cytokines had no effect on transcription from the rous sarcoma virus promoter or the SV40 virus early promoter. Protein binding assays indicate that TNF and gamma-INF did not effect the ability of the ME1a2 factor to bind to its site but did significantly repress E2F factor binding to its DNA sequence.

Cyclin A levels, the duration of S phase and sensitivity to a chemotherapeutic agent are altered in fibroblasts cultured on a fibronectin matrix.

Culture of murine embryonic fibroblasts, but not vascular smooth muscle cells, on a fibronectin matrix significantly shortens their transit time through the S phase of the cell cycle. This shortening corresponds to an increase in both cyclin A protein levels and active cyclin A/cdk2 complex. The increase in cyclin A protein appears due to a translational/post-translational mechanism since there is no increase in cyclin A mRNA following culture of the cells on fibronectin. Treatment of cells cultured on fibronectin with a short pulse of the S phase chemotherapeutic agent camptothecin, resulted in a relative protection from cell death when compared to cells cultured on tissue culture plastic. Thus, while the cells have increased rate of transit through S phase fibronectin-mediated signaling protects the cells from S phase mediated apoptosis. In addition, fibroblasts constitutively expressing a mutant E2F1 transcription factor (E2F1d87) have a lengthened S phase, due to a truncation of the cyclin A/cdk2 binding domain. Culture of these mutant- expressing cells on fibronectin did not shorten their S phase duration in spite of the fact that cyclin A levels and active cyclin A/cdk2 complex were significantly elevated. Thus, although the fibronectin signaling mechanisms culminating in elevated cyclin A were intact in these mutant E2F1 expressing cells, they were insensitive to the effects of this elevated cyclin A. The effect of the mutant E2F1d87 on slowing transit through S phase appears dominant over the effect of elevated cyclin A.

Primary malignant melanoma of the uterine cervix.

A case of primary malignant melanoma of the cervix is presented that was treated by radical pelvic surgery. The cytology, histology and electron microscopy of the lesion are presented with a brief clinical discussion.

Kaposi's sarcoma of the vulva: a case report and brief review.

A report of a case of Kaposi's sarcoma of the vulva is presented. A complete response of the tumor was obtained after 1 treatment cycle using a triple-drug chemotherapy regimen of actinomycin D, vincristine, and 5-(3,3-dimethyl-1-trazeno)imidazole-4-carboxamide (DTIC). Death occurred due to severe erythema multiforme, which was indirectly related to a complication of the chemotherapy.

A new technique for vulvar skin grafting.

A new and easy technique for handling and applying a split-thickness skin graft to the vulva is described. The use of petrolatum gauze backing and application with the skin stapler, as well as postoperative management, are discussed.

Bleomycin-mitomycin C in advanced squamous cell carcinoma of the female genital tract.

Bleomycin and mitomycin C have been reported to be active against advanced squamous cell carcinoma of the uterine cervix. Miyamoto et al reported a total response rate of 93% and complete response in 80% of the first 15 patients treated by sequential combination of bleomycin and mitomycin C. Updated results by Miyamoto reveal an 88.3% total response rate with a 65.3% complete response. However, others have reported poor results using the protocol outlined by Miyamoto. Using this protocol, 11 patients with advanced squamous cell carcinoma were treated at the authors' institution. There were no complete responses: one patient with partial response (9%), four with stable disease (36%), and six (55%) with progressive disease.

The spatial resolution performance of a time-resolved optical imaging system using temporal extrapolation.

Optical imaging methods are being explored as a potential means of screening for breast cancer. Previous investigations of time-resolved imaging techniques have suggested that due to the lack of photons with sufficiently small pathlengths, the spatial resolution achievable through a human breast would be unlikely to be better than a centimeter. Experimental results presented here indicate, however, that higher resolution may be achieved by extrapolating the measured temporal distribution of transmitted photons. This is performed using a least-squares fit between data and an analytic model of photon transport. The spatial resolution of a time-resolved imaging system was evaluated by measuring the edge response produced by an opaque mask embedded in the center of a 51-mm-thick, very highly scattering medium. The limiting spatial resolution was improved from about 13 mm to about 5 mm.