Immunogenicity and Safety of a Purified Vero Rabies Vaccine-Serum Free, Compared With 2 Licensed Vaccines, in a Simulated Rabies Post-Exposure Regimen in Healthy Adults in France: A Randomized, Controlled, Phase 3 Trial.

BACKGROUND: A next-generation Vero cell rabies vaccine (PVRV-NG2) was developed using the same Pitman-Moore strain as in the licensed purified Vero cell vaccine (PVRV; Verorab) and the human diploid cell vaccine (HDCV; Imovax Rabies®). METHODS: This dual-center, modified, double-blind, phase 3 study evaluated the immunogenic non-inferiority and safety of PVRV-NG2 with and without concomitant intramuscular human rabies immunoglobulin (HRIG) versus PVRV + HRIG and HDCV + HRIG in a simulated post-exposure prophylaxis (PEP) regimen. Healthy adults ≥18 years old (N = 640) were randomized 3:1:1:1 to PVRV-NG2 + HRIG, PVRV + HRIG, HDCV + HRIG, or PVRV-NG2 alone (administered as single vaccine injections on days [D] 0, D3, D7, D14, and 28, with HRIG on D0 in applicable groups). Rabies virus neutralizing antibodies (RVNA) titers were assessed pre- (D0) and post-vaccination (D14, D28, and D42) using the rapid fluorescent focus inhibition test. Non-inferiority, based on the proportion of participants achieving RVNA titers ≥0.5 IU/mL (primary objective), was demonstrated if the lower limit of the 95% CI of the difference in proportions between PVRV-NG2 + HRIG and PVRV + HRIG/HDCV + HRIG was >-5% at D28. Safety was assessed up to 6 months after the last injection. RESULTS: Non-inferiority of PVRV-NG2 + HRIG compared with PVRV + HRIG and HDCV + HRIG was demonstrated. Nearly all participants (99.6%, PVRV-NG2 + HRIG; 100%, PVRV + HRIG; 98.7%, HDCV + HRIG; 100%, PVRV-NG2 alone) achieved RVNA titers ≥0.5 IU/mL at D28. Geometric mean titers were similar between groups with concomitant HRIG administration at all time points. Safety profiles were similar between PVRV-NG2 and comparator vaccines. CONCLUSIONS: In a simulated PEP setting, PVRV-NG2 + HRIG showed comparable immunogenicity and safety to current standard-of-care vaccines. CLINICAL TRIALS REGISTRATION: NCT03965962.

Current vaccination strategy against Piscirickettsia salmonis in Chile based only on the EM-90 genogroup shows incomplete cross-protection for the LF-89 genogroup.

Piscirickettsia salmonis, the primary bacterial disease in Chilean salmon farming, necessitates a constant refinement of control strategies. This study hypothesized that the current vaccination strategy for SRS control in the Chilean Atlantic salmon aquaculture industry, which has been in place since 2017 (ALPHA JECT® 5.1 plus LiVac®), solely relies on vaccines formulated with the EM-90 genogroup of P. salmonis (PS-EM-90), triggering a partial cross-immunity response in fish infected with the LF-89 genogroup (PS-LF-89). Relative Percent Survival (RPS) and cell-mediated immune (CMI) response were evaluated in Atlantic salmon post-smolts vaccinated with the standard vaccination strategy but challenged with both PS-EM-90 and PS-LF-89, in addition to other vaccination strategies considering primo vaccination and booster with other commercial vaccines and the possible enhancing effects of the combination with a natural immunomodulator (PAQ-Xtract®) administered orally. The intraperitoneal (I.P.) challenge was performed after 2395°-days (DD) after the start of the immunostimulant delivery, 1905 DD after the primo vaccination, and 1455 DD after the booster vaccination. Unvaccinated fish showed 73.6 and 41.7 % mortality when challenged with PS-EM-90 and PS-LF-89, respectively. Fish infected with PS-LF-89 died significantly faster (21 days post-infection, dpi) than fish challenged with PS-EM-90 (28 dpi) (p = 0.0043) and had a higher probability of death (0.4626) than fish challenged with PS-EM-90. RPS had a significant positive correlation with the PS-EM-90 load of the P. salmonis genogroup (r = 0.540, p < 0.01) but not with the PS-LF-89 load (r = 0.155, p > 0.05). This demonstrated that the immunization strategies were more effective in lowering PS-EM-90 loads, resulting in higher survival rates in fish challenged with PS-EM-90. The current industry vaccination strategy recorded a 100 % RPS when fish were challenged with PS-EM-90, but the RPS dropped significantly to 77 % when fish were challenged with PS-LF-89, meaning that the strategy did not show complete cross-protection. But after adding PAQ-Xtract®, the RPS improved from 77 % to 92 % in fish that were vaccinated with the standard method but then challenged with PS-LF-89. The most effective vaccination strategy was based on LiVac® as primo vaccination and ALPHA JECT® 5.1 plus LiVac® as booster vaccination, with or without PAQ-Xtract®, in both PS-EM-90 (100 %) and PS-LF-89 (96 %) challenged fish. The serum concentration of anti-P. salmonis IgM did not show a correlation with the protection of immunization strategies expressed in survival. Low serum IL-12 and high serum IFNγ concentrations showed a correlation with higher bacterial loads and lower survival. Aggregate analysis showed a significant correlation between higher numbers of CD8+ cells in the head-kidney, higher fish survival, and a lower bacterial load. The immunization strategies were safe for fish and induced only mild microscopic lesions in the gut. Taken together, our results help to better understand the biological interaction between P. salmonis and post-smolt vaccinated Atlantic salmon to deepen the knowledge on vaccine-induced protection, CMI immune response, and cross-immunity applied to improve the current immunization strategy for SRS control in the Chilean salmon industry.

Specific microRNA Profile Associated with Inflammation and Lipid Metabolism for Stratifying Allergic Asthma Severity.

The mechanisms underlying severe allergic asthma are complex and unknown, meaning it is a challenge to provide the most appropriate treatment. This study aimed to identify novel biomarkers for stratifying allergic asthmatic patients according to severity, and to uncover the biological mechanisms that lead to the development of the severe uncontrolled phenotype. By using miRNA PCR panels, we analyzed the expression of 752 miRNAs in serum samples from control subjects (n = 15) and mild (n = 11) and severe uncontrolled (n = 10) allergic asthmatic patients. We identified 40 differentially expressed miRNAs between severe uncontrolled and mild allergic asthmatic patients. Functional enrichment analysis revealed signatures related to inflammation, angiogenesis, lipid metabolism and mRNA regulation. A random forest classifier trained with DE miRNAs achieved a high accuracy of 97% for severe uncontrolled patient stratification. Validation of the identified biomarkers was performed on a subset of allergic asthmatic patients from the CAMP cohort at Brigham and Women's Hospital, Harvard Medical School. Four of these miRNAs (hsa-miR-99b-5p, hsa-miR-451a, hsa-miR-326 and hsa-miR-505-3p) were validated, pointing towards their potential as biomarkers for stratifying allergic asthmatic patients by severity and providing insights into severe uncontrolled asthma molecular pathways.

CRISPRi-induced transcriptional regulation of IAH1 gene and its influence on volatile compounds profile in Kluyveromyces marxianus DU3.

Mezcal is a traditional Mexican distilled beverage, known for its marked organoleptic profile, which is influenced by several factors, such as the fermentation process, where a wide variety of microorganisms are present. Kluyveromyces marxianus is one of the main yeasts isolated from mezcal fermentations and has been associated with ester synthesis, contributing to the flavors and aromas of the beverage. In this study, we employed CRISPR interference (CRISPRi) technology, using dCas9 fused to the Mxi1 repressor factor domain, to down-regulate the expression of the IAH1 gene, encoding for an isoamyl acetate-hydrolyzing esterase, in K. marxianus strain DU3. The constructed CRISPRi plasmid successfully targeted the IAH1 gene, allowing for specific gene expression modulation. Through gene expression analysis, we assessed the impact of IAH1 down-regulation on the metabolic profile of volatile compounds. We also measured the expression of other genes involved in volatile compound biosynthesis, including ATF1, EAT1, ADH1, and ZWF1 by RT-qPCR. Results demonstrated successful down-regulation of IAH1 expression in K. marxianus strain DU3 using the CRISPRi system. The modulation of IAH1 gene expression resulted in alterations in the production of volatile compounds, specifically ethyl acetate, which are important contributors to the beverage's aroma. Changes in the expression levels of other genes involved in ester biosynthesis, suggesting that the knockdown of IAH1 may generate intracellular alterations in the balance of these metabolites, triggering a regulatory response. The application of CRISPRi technology in K. marxianus opens the possibility of targeted modulation of gene expression, metabolic engineering strategies, and synthetic biology in this yeast strain.

Distinct rates and patterns of spread of the major HIV-1 subtypes in Central and East Africa.

Since the ignition of the HIV-1 group M pandemic in the beginning of the 20th century, group M lineages have spread heterogeneously throughout the world. Subtype C spread rapidly through sub-Saharan Africa and is currently the dominant HIV lineage worldwide. Yet the epidemiological and evolutionary circumstances that contributed to its epidemiological expansion remain poorly understood. Here, we analyse 346 novel pol sequences from the DRC to compare the evolutionary dynamics of the main HIV-1 lineages, subtypes A1, C and D. Our results place the origins of subtype C in the 1950s in Mbuji-Mayi, the mining city of southern DRC, while subtypes A1 and D emerged in the capital city of Kinshasa, and subtypes H and J in the less accessible port city of Matadi. Following a 15-year period of local transmission in southern DRC, we find that subtype C spread at least three-fold faster than other subtypes circulating in Central and East Africa. In conclusion, our results shed light on the origins of HIV-1 main lineages and suggest that socio-historical rather than evolutionary factors may have determined the epidemiological fate of subtype C in sub-Saharan Africa.

Cost-effectiveness of longitudinal surveillance for Piscirickettsia salmonis using qPCR in Atlantic salmon farms (Salmo salar) in Chile.

Costs of diagnostic testing including sample collection, sampling frequency and sample size are an important consideration in the evaluation of the economic feasibility of alternative surveillance strategies for detection of infectious diseases in aquatic animals. In Chile, Piscirickettsia salmonis is the primary reason for antibiotic treatments in farmed Atlantic salmon. In 2012, a surveillance and control programme for piscirickettsiosis was established with an overall goal of reducing antibiotic use. The present study estimated the cost-effectiveness of different sampling frequencies and sample sizes to achieve at least 95% confidence of early detection of P. salmonis at the netpen and farm levels using a validated qPCR test. We developed a stochastic model that incorporated variability in test accuracy, within-pen prevalence and sampling costs. Our findings indicated that the current piscirickettsiosis surveillance programme based on risk-based sampling of five moribund or dead fish from 2 to 3 netpens is cost-effective and gives a high probability of detection of P. salmonis in Atlantic salmon farms in Chile at both the netpen and farm levels. Results from this study should incentivize salmon farmers to establish cost-effective strategies for early detection of P. salmonis infection and the application of this approach to other highly infectious diseases.

Effect of Lupinus rotundiflorus gamma conglutin treatment on JNK1 gene expression and protein activation in a rat model of type 2 diabetes.

CONTEXT: Gamma conglutin (Cγ) from lupine species represents a potential complementary treatment for type 2 diabetes mellitus (T2DM) because of its hypoglycaemic effect. However, its underlying mechanism of action is not fully known. OBJECTIVE: To evaluate whether Cγ from Lupinus rotundiflorus M. E. Jones (Fabaceae) modulates c-Jun N-terminal kinase 1 (JNK1) expression and activation in a T2DM rat model. MATERIALS AND METHODS: Gamma conglutin isolated from L. rotundiflorus seeds was characterized by SDS-PAGE. Fifteen Wistar rats with streptozotocin-induced T2DM (HG) were randomized into three groups (n = 5): vehicle administration (HG-Ctrl), oral treatment with Cγ (120 mg/kg/day) (HG-Lr) for one week, and treatment with metformin (300 mg/kg/day) (HG-Met); a healthy group (Ctrl, n = 5) was included as control. The levels of glucose and biomarkers of renal and hepatic function were measured pre- and post-treatment. Hepatic Jnk1 expression and phosphorylation of JNK1 were evaluated by qRT-PCR and western blot, respectively. RESULTS: Oral treatment with either Cγ or metformin reduced serum glucose level to 86.30 and 74.80 mg/dL, respectively (p ˂ 0.05), from the basal levels. Jnk1 expression was 0.65- and 0.54-fold lower (p ˂ 0.05) in the HG-Lr and HG-Met groups, respectively, than in HG-Ctrl. Treatment with Cγ decreased JNK1 phosphorylation. However, Cγ did not change the levels of kidney and liver biomarkers. DISCUSSION AND CONCLUSIONS: Treatment with Cγ from L. rotundiflorus inhibited Jnk1 expression, in vivo, suggesting JNK1 as a potential therapeutic target in diabetes and revealing one mechanism underlying the hypoglycaemic effect of lupine Cγ. Nevertheless, further studies are required.

Bayesian estimation of diagnostic sensitivity and specificity of a qPCR and a bacteriological culture method for Piscirickettsia salmonis in farmed Atlantic salmon (Salmo salar L.) in Chile.

Early detection of piscirickettsiosis is an important purpose of government- and industry-based surveillance for the disease in Atlantic salmon farms in Chile. Real-time qPCRs are currently used for surveillance because bacterial isolation is inadequately sensitive or rapid enough for routine use. Since no perfect tests exist, we used Bayesian latent class models to estimate diagnostic sensitivity (DSe) and specificity (DSp) of qPCR and culture using separate two-test, single-population models for three farms (n = 148, 151, 44). Informative priors were used for DSp (culture (beta(999,1); qPCR (beta(98,2)), and flat priors (beta 1,1) for DSe and prevalence. Models were run for liver and kidney tissues combined and separately, based on the presence of selected gross-pathological signs. Across all models, qPCR DSe was 5- to 30-fold greater than for culture. Combined-tissue qPCR median DSe was highest in Farm 3 (sampled during P. salmonis outbreak (DSe = 97.6%)) versus Farm 1 (DSe = 85.6%) or Farm 2 (DSe = 83.5%), both sampled before clinical disease. Median DSe of qPCR was similar for liver and kidney, but higher when gross-pathological signs were evident at necropsy. High DSe and DSp and rapid turnaround-time indicate that the qPCR is fit for surveillance programmes and diagnosis during an outbreak. Targeted testing of salmon with gross-pathological signs can enhance DSe.

Tracing the Impact of Public Health Interventions on HIV-1 Transmission in Portugal Using Molecular Epidemiology.

BACKGROUND: Estimation of temporal changes in human immunodeficiency virus (HIV) transmission patterns can help to elucidate the impact of preventive strategies and public health policies. METHODS: Portuguese HIV-1 subtype B and G pol genetic sequences were appended to global reference data sets to identify country-specific transmission clades. Bayesian birth-death models were used to estimate subtype-specific effective reproductive numbers (Re). Discrete trait analysis (DTA) was used to quantify mixing among transmission groups. RESULTS: We identified 5 subtype B Portuguese clades (26-79 sequences) and a large monophyletic subtype G Portuguese clade (236 sequences). We estimated that major shifts in HIV-1 transmission occurred around 1999 (95% Bayesian credible interval [BCI], 1998-2000) and 2000 (95% BCI, 1998-2001) for subtypes B and G, respectively. For subtype B, Re dropped from 1.91 (95% BCI, 1.73-2.09) to 0.62 (95% BCI,.52-.72). For subtype G, Re decreased from 1.49 (95% BCI, 1.39-1.59) to 0.72 (95% BCI, .63-.8). The DTA suggests that people who inject drugs (PWID) and heterosexuals were the source of most (>80%) virus lineage transitions for subtypes G and B, respectively. CONCLUSIONS: The estimated declines in Re coincide with the introduction of highly active antiretroviral therapy and the scale-up of harm reduction for PWID. Inferred transmission events across transmission groups emphasize the importance of prevention efforts for bridging populations.

Gene expression associated with immune response in Atlantic salmon head-kidney vaccinated with inactivated whole-cell bacterin of Piscirickettsia salmonis and pathogenic isolates.

Piscirickettsiosis is the most challenging disease present in the Chilean salmon industry. The aim of this study was to describe the expression of genes associated with immune response of Atlantic salmon intraperitoneally infected with LF-89 and EM-90 Piscirickettsia salmonis and vaccinated with inactivated whole-cell bacterin of P. salmonis. The fish infected with PS-LF-89 showed an anti-inflammatory response, whereas this finding was not observed in the PS-EM-90-infected fish and vaccinated fish. Fish infected with both P. salmonis isolates showed mhc1-mhc2, cd4-cd8b and igm overexpression, suggesting that P. salmonis promotes a T CD4+ and T CD8+ cell response and a humoral immune response. The vaccinated-fish exhibited mhc1, mhc2 and cd4 overexpression but a significant downregulation of cd8b and igm, suggesting that the vaccine supported the CD4+ T-cell response but did not induce an immune response mediated by CD8+ T cells or a humoral response. In conclusion, the expression pattern of genes related to the humoral and cell-mediated adaptive immune response showed upregulation in fish infected with P. salmonis and down-regulation in vaccinated fish. The results of this study contribute to our understanding of the immune response against P. salmonis and can be used in the optimization of SRS prevention and control measures.

First Latin American clinical practice guidelines for the treatment of systemic lupus erythematosus: Latin American Group for the Study of Lupus (GLADEL, Grupo Latino Americano de Estudio del Lupus)-Pan-American League of Associations of Rheumatology (PANLAR).

Systemic lupus erythematosus (SLE), a complex and heterogeneous autoimmune disease, represents a significant challenge for both diagnosis and treatment. Patients with SLE in Latin America face special problems that should be considered when therapeutic guidelines are developed. The objective of the study is to develop clinical practice guidelines for Latin American patients with lupus. Two independent teams (rheumatologists with experience in lupus management and methodologists) had an initial meeting in Panama City, Panama, in April 2016. They selected a list of questions for the clinical problems most commonly seen in Latin American patients with SLE. These were addressed with the best available evidence and summarised in a standardised format following the Grading of Recommendations Assessment, Development and Evaluation approach. All preliminary findings were discussed in a second face-to-face meeting in Washington, DC, in November 2016. As a result, nine organ/system sections are presented with the main findings; an 'overarching' treatment approach was added. Special emphasis was made on regional implementation issues. Best pharmacologic options were examined for musculoskeletal, mucocutaneous, kidney, cardiac, pulmonary, neuropsychiatric, haematological manifestations and the antiphospholipid syndrome. The roles of main therapeutic options (ie, glucocorticoids, antimalarials, immunosuppressant agents, therapeutic plasma exchange, belimumab, rituximab, abatacept, low-dose aspirin and anticoagulants) were summarised in each section. In all cases, benefits and harms, certainty of the evidence, values and preferences, feasibility, acceptability and equity issues were considered to produce a recommendation with special focus on ethnic and socioeconomic aspects. Guidelines for Latin American patients with lupus have been developed and could be used in similar settings.

Association of HIV status with infection by multiple HPV types.

OBJECTIVES: To identify the clinical and demographic characteristics of HIV-positive and HIV-negative women infected by multiple HPV types. METHODS: 1399 women participated in the study (240 HIV-positive and 1159 HIV-negative women). Samples were provided for Pap tests and for HPV detection and typing by PCR. Data were collected on HPV infection, frequency of multiple infection, and HPV type distribution. Odds ratios were reported from logistic regression models. RESULTS: Compared with HIV-negative women, HIV-positive women had higher frequencies of cervical abnormality (30% vs. 20.8%), higher HPV prevalence (68.3% vs. 51.3%) and were more commonly infected with multiple HPV types (78.7% vs. 44.3%). HPV-16 was the most common type detected in the study population, with other types showing variable associations with HIV status. Positive associations were observed between infection by multiple HPV types and HIV status, cervical abnormality and having had more than three pregnancies. The odds of multiple infection by HPV types were higher in HIV-positive women who used an intrauterine device, who had a history of abortions and who had HIV viral loads >100 000 copies/ml, whilst the odds were lower in women with >500 CD4 cells/mm3 . CONCLUSIONS: HIV immunosuppression favours infection by multiple high-risk HPV types, mainly in women affected by low-grade squamous intraepithelial lesions. Antiretroviral therapy had no effect on infection by multiple HPV types. Risk factors related to progressive damage to the cervix were positively associated with infection by multiple HPV types in women living with HIV.

Implications of hepatitis C virus subtype 1a migration patterns for virus genetic sequencing policies in Italy.

BACKGROUND: In-depth phylogeographic analysis can reveal migration patterns relevant for public health planning. Here, as a model, we focused on the provenance, in the current Italian HCV subtype 1a epidemic, of the NS3 resistance-associated variant (RAV) Q80K, known to interfere with the action of NS3/4A protease inhibitor simeprevir. HCV1a migration patterns were analysed using Bayesian phylodynamic tools, capitalising on newly generated and publicly available time and geo-referenced NS3 encoding virus genetic sequence data. RESULTS: Our results showed that both immigration and local circulation fuel the current Italian HCV1a epidemic. The United States and European continental lineages dominate import into Italy, with the latter taking the lead from the 1970s onwards. Since similar migration patterns were found for Q80K and other lineages, no clear differentiation of the risk for failing simeprevir can be made between patients based on their migration and travel history. Importantly, since HCV only occasionally recombines, these results are readily transferable to the genetic sequencing policy concerning NS5A RAVs. CONCLUSIONS: The patient migration and travel history cannot be used to target only part of the HCV1a infected population for drug resistance testing before start of antiviral therapy. Consequently, it may be cost-effective to expand genotyping efforts to all HCV1a infected patients eligible for simeprevir-based therapies.

An integrated map of HIV genome-wide variation from a population perspective.

BACKGROUND: The HIV pandemic is characterized by extensive genetic variability, which has challenged the development of HIV drugs and vaccines. Although HIV genomes have been classified into different types, groups, subtypes and recombinants, a comprehensive study that maps HIV genome-wide diversity at the population level is still lacking to date. This study aims to characterize HIV genomic diversity in large-scale sequence populations, and to identify driving factors that shape HIV genome diversity. RESULTS: A total of 2996 full-length genomic sequences from 1705 patients infected with 16 major HIV groups, subtypes and circulating recombinant forms (CRFs) were analyzed along with structural, immunological and peptide inhibitor information. Average nucleotide diversity of HIV genomes was almost 50% between HIV-1 and HIV-2 types, 37.5% between HIV-1 groups, 14.7% between HIV-1 subtypes, 8.2% within individual HIV-1 subtypes and less than 1% within single patients. Along the HIV genome, diversity patterns and compositions of nucleotides and amino acids were highly similar across different groups, subtypes and CRFs. Current HIV-derived peptide inhibitors were predominantly derived from conserved, solvent accessible and intrinsically ordered structures in the HIV-1 subtype B genome. We identified these conserved regions in Capsid, Nucleocapsid, Protease, Integrase, Reverse transcriptase, Vpr and the GP41 N terminus as potential drug targets. In the analysis of factors that impact HIV-1 genomic diversity, we focused on protein multimerization, immunological constraints and HIV-human protein interactions. We found that amino acid diversity in monomeric proteins was higher than in multimeric proteins, and diversified positions were preferably located within human CD4 T cell and antibody epitopes. Moreover, intrinsic disorder regions in HIV-1 proteins coincided with high levels of amino acid diversity, facilitating a large number of interactions between HIV-1 and human proteins. CONCLUSIONS: This first large-scale analysis provided a detailed mapping of HIV genomic diversity and highlighted drug-target regions conserved across different groups, subtypes and CRFs. Our findings suggest that, in addition to the impact of protein multimerization and immune selective pressure on HIV-1 diversity, HIV-human protein interactions are facilitated by high variability within intrinsically disordered structures.

Human papillomavirus detection in women with and without human immunodeficiency virus infection in Colombia.

BACKGROUND: HIV infection leads to a decreasing immune response, thereby facilitating the appearance of other infections, one of the most important ones being HPV. However, studies are needed for determining associations between immunodeficiency caused by HIV and/or the presence of HPV during the course of cervical lesions and their degree of malignancy. This study describes the cytological findings revealed by the Papanicolaou test, laboratory characteristics and HPV molecular profile in women with and without HIV infection. METHODS: A total of 216 HIV-positive and 1,159 HIV-negative women were invited to participate in the study; PCR was used for the molecular detection of HPV in cervical samples. Statistical analysis (such as percentages, Chi-square test and Fisher's exact test when applicable) determined human papillomavirus (HPV) infection frequency (single and multiple) and the distribution of six types of high-risk-HPV in women with and without HIV infection. Likewise, a logistic regression model was run to evaluate the relationship between HIV-HPV infection and different risk factors. RESULTS: An association was found between the frequency of HPV infection and infection involving 2 or more HPV types (also known as multiple HPV infection) in HIV-positive women (69.0% and 54.2%, respectively); such frequency was greater than that found in HIV-negative women (44.3% and 22.7%, respectively). Statistically significant differences were observed between both groups (p = 0.001) regarding HPV presence (both in infection and multiple HPV infection). HPV-16 was the most prevalent type in the population being studied (p = 0.001); other viral types had variable distribution in both groups (HIV-positive and HIV-negative). HPV detection was associated with <500 cell/mm(3) CD4-count (p = 0.004) and higher HIV-viral-load (p = 0.001). HPV-DNA detection, <200 cell/mm(3) CD4-count (p = 0.001), and higher HIV-viral-load (p = 0.001) were associated with abnormal cytological findings. CONCLUSIONS: The HIV-1 positive population in this study had high multiple HPV infection prevalence. The results for this population group also suggested a greater association between HPV-DNA presence and cytological findings. HPV detection, together with low CD4 count, could represent useful tools for identifying HIV-positive women at risk of developing cervical lesions.

"Trust Women": Characteristics of and learnings from patients of a Shield Law medication abortion practice in the United States.

INTRODUCTION: The 2022 Massachusetts Shield Law protects telemedicine providers who care for abortion seekers in other states from criminal, civil, and licensure penalties. In this article we explore the characteristics of patients of The Massachusetts Medication Abortion Access Project (The MAP). METHODS: The MAP is an asynchronous telemedicine service that offers mifepristone/misoprostol to abortion seekers in all 50 states who are at or under 11 weeks pregnancy gestation on initial intake. The MAP charges USD250 using a pay-what-you-can model. We analyzed medical questionnaires and payments submitted by patients who received care from The MAP during its first 6 months of operations using descriptive statistics and for content and themes. RESULTS: From October 1, 2023-March 31, 2024, 1994 patients accessed care through The MAP. Almost all (n = 1973, 99%) identified as women/girls and about half (n = 984, 49%) were aged 20-29. The MAP cared for patients in 45 states; 84% (n = 1672) of these patients received pills in abortion ban or restricted southern states. Patients paid USD134.50 on average; 29% (n = 577) paid USD25 or less. Nearly two-thirds (n = 1293, 65%) received subsidized care; financial hardship featured prominently in patient comments. DISCUSSION: Considerable demand exists for medication abortion care from Shield Law providers. The MAP demonstrates that providers can trust women and other pregnancy capable people to decide for themselves whether to obtain medication abortion pills by mail and to pay what they can afford without being required to justify their need. Identifying ways to support Shield Law provision and further subsidize abortion care are needed.

Pseudomonas aeruginosa surface motility and invasion into competing communities enhance interspecies antagonism.

Chronic polymicrobial infections involving Pseudomonas aeruginosa and Staphylococcus aureus are prevalent, difficult to eradicate, and associated with poor health outcomes. Therefore, understanding interactions between these pathogens is important to inform improved treatment development. We previously demonstrated that P. aeruginosa is attracted to S. aureus using type IV pili (TFP)-mediated chemotaxis, but the impact of attraction on S. aureus growth and physiology remained unknown. Using live single-cell confocal imaging to visualize microcolony structure, spatial organization, and survival of S. aureus during coculture, we found that interspecies chemotaxis provides P. aeruginosa a competitive advantage by promoting invasion into and disruption of S. aureus microcolonies. This behavior renders S. aureus susceptible to P. aeruginosa antimicrobials. Conversely, in the absence of TFP motility, P. aeruginosa cells exhibit reduced invasion of S. aureus colonies. Instead, P. aeruginosa builds a cellular barrier adjacent to S. aureus and secretes diffusible, bacteriostatic antimicrobials like 2-heptyl-4-hydroxyquinoline-N-oxide (HQNO) into the S. aureus colonies. Reduced invasion leads to the formation of denser and thicker S. aureus colonies with increased HQNO-mediated lactic acid fermentation, a physiological change that could complicate treatment strategies. Finally, we show that P. aeruginosa motility modifications of spatial structure enhance competition against S. aureus. Overall, these studies expand our understanding of how P. aeruginosa TFP-mediated interspecies chemotaxis facilitates polymicrobial interactions, highlighting the importance of spatial positioning in mixed-species communities. IMPORTANCE: The polymicrobial nature of many chronic infections makes their eradication challenging. Particularly, coisolation of Pseudomonas aeruginosa and Staphylococcus aureus from airways of people with cystic fibrosis and chronic wound infections is common and associated with severe clinical outcomes. The complex interplay between these pathogens is not fully understood, highlighting the need for continued research to improve management of chronic infections. Our study unveils that P. aeruginosa is attracted to S. aureus, invades into neighboring colonies, and secretes anti-staphylococcal factors into the interior of the colony. Upon inhibition of P. aeruginosa motility and thus invasion, S. aureus colony architecture changes dramatically, whereby S. aureus is protected from P. aeruginosa antagonism and responds through physiological alterations that may further hamper treatment. These studies reinforce accumulating evidence that spatial structuring can dictate community resilience and reveal that motility and chemotaxis are critical drivers of interspecies competition.

Pseudomonas aeruginosa surface motility and invasion into competing communities enhances interspecies antagonism.

Chronic polymicrobial infections involving Pseudomonas aeruginosa and Staphylococcus aureus are prevalent, difficult to eradicate, and associated with poor health outcomes. Therefore, understanding interactions between these pathogens is important to inform improved treatment development. We previously demonstrated that P. aeruginosa is attracted to S. aureus using type IV pili-mediated chemotaxis, but the impact of attraction on S. aureus growth and physiology remained unknown. Using live single-cell confocal imaging to visualize microcolony structure, spatial organization, and survival of S. aureus during coculture, we found that interspecies chemotaxis provides P. aeruginosa a competitive advantage by promoting invasion into and disruption of S. aureus microcolonies. This behavior renders S. aureus susceptible to P. aeruginosa antimicrobials. Conversely, in the absence of type IV pilus motility, P. aeruginosa cells exhibit reduced invasion of S. aureus colonies. Instead, P. aeruginosa builds a cellular barrier adjacent to S. aureus and secretes diffusible, bacteriostatic antimicrobials like 2-heptyl-4-hydroxyquinoline-N-oxide (HQNO) into the S. aureus colonies. P. aeruginosa reduced invasion leads to the formation of denser and thicker S. aureus colonies with significantly increased HQNO-mediated lactic acid fermentation, a physiological change that could complicate the effective treatment of infections. Finally, we show that P. aeruginosa motility modifications of spatial structure enhance competition against S. aureus. Overall, these studies build on our understanding of how P. aeruginosa type IV pili-mediated interspecies chemotaxis mediates polymicrobial interactions, highlighting the importance of spatial positioning in mixed-species communities.

Identification of Black Cumin (Nigella sativa) MicroRNAs by Next-Generation Sequencing and Their Implications in Secondary Metabolite Biosynthesis.

Secondary metabolites are bioactive compounds believed to contribute to the pharmacological properties of plants. MicroRNAs (miRNAs) are small non-coding RNA molecules involved in post-transcriptional regulation and are thought to play an important role in regulating secondary metabolism biosynthesis. Nevertheless, the extent of miRNA involvement in secondary metabolism remains minimal. Nigella sativa (black cumin/black seed) is a popular medicinal and culinary plant known for its pharmaceutical properties; however, its genomic information is scarce. In this study, next-generation sequencing (NGS) technology was employed to obtain the miRNA profile of N. sativa, and their involvement in secondary metabolite biosynthesis was explored. A total of 25,139,003 unique reads ranging from 16 to 40 nucleotides were attained, out of which 240 conserved and 34 novel miRNAs were identified. Moreover, 6083 potential target genes were recognized in this study. Several conserved and novel black cumin miRNAs were found to target enzymes involved in the terpenoid, diterpenoid, phenylpropanoid, carotenoid, flavonoid, steroid, and ubiquinone biosynthetic pathways, among others, for example, beta-carotene 3-hydroxylase, gibberellin 3 beta-dioxygenase, trimethyltridecatetraene synthase, carboxylic ester hydrolases, acetyl-CoA C-acetyltransferase, isoprene synthase, peroxidase, shikimate O-hydroxycinnamoyltransferase, etc. Furthermore, sequencing data were validated through qPCR by checking the relative expression of eleven randomly selected conserved and novel miRNAs (nsa-miR164d, nsa-miR166a, nsa-miR167b, nsa-miR171a, nsa-miR390b, nsa-miR396, nsa-miR159a, nsa-miRN1, nsa-miRN29, nsa-miRN32, and nsa-miRN34) and their expression patterns were found to be corroborated with the sequencing data. We anticipate that this work will assist in clarifying the implications of miRNAs in plant secondary metabolism and aid in the generation of artificial miRNA-based strategies to overproduce highly valuable secondary metabolites from N. sativa.