Zoonotic transmission of diphtheria toxin-producing Corynebacterium ulcerans.

BACKGROUND: Diphtheria caused by toxin-producing Corynebacterium ulcerans is a re-emerging human disease that can cause local and systemic sequelae. In Australia, toxigenic diphtheria is a rare notifiable communicable disease, due to high-vaccination coverage. The public health management of cutaneous cases of toxigenic C. ulcerans varies between jurisdictions, as opposed to the more uniform public health response to toxigenic Corynebacterium diphtheriae presenting as respiratory or laryngeal diphtheria. AIM: To report a case of zoonotically acquired C. ulcerans, review evidence on the zoonotic reservoir and reported transmission events, and examine public health guidelines for the management of human and animal contacts. METHODS AND RESULTS: In this case report, we detail our case investigation, treatment and public health management, including contact tracing and an approach to animal testing. We successfully identified companion canines as probable sources for the human case, with WGS confirming the link. The zoonotic disease link of C. ulcerans to domestic and agricultural animals is established in the literature; however, the management of animal contacts in human cases is inconsistent with jurisdictional or national guidelines. CONCLUSIONS: While a rare disease, a consistent approach to public health management is warranted to systematically elucidate the disease source and improve understanding of transmission.

Gene dysregulation in acute HIV-1 infection - early transcriptomic analysis reveals the crucial biological functions affected.

Introduction: Transcriptomic analyses from early human immunodeficiency virus (HIV) infection have the potential to reveal how HIV causes widespread and lasting damage to biological functions, especially in the immune system. Previous studies have been limited by difficulties in obtaining early specimens. Methods: A hospital symptom-based screening approach was applied in a rural Mozambican setting to enrol patients with suspected acute HIV infection (Fiebig stage I-IV). Blood samples were collected from all those recruited, so that acute cases and contemporaneously recruited, uninfected controls were included. PBMC were isolated and sequenced using RNA-seq. Sample cellular composition was estimated from gene expression data. Differential gene expression analysis was completed, and correlations were determined between viral load and differential gene expression. Biological implications were examined using Cytoscape, gene set enrichment analysis, and enrichment mapping. Results: Twenty-nine HIV infected subjects one month from presentation and 46 uninfected controls were included in this study. Subjects with acute HIV infection demonstrated profound gene dysregulation, with 6131 (almost 13% of the genome mapped in this study) significantly differentially expressed. Viral load was correlated with 1.6% of dysregulated genes, in particular, highly upregulated genes involved in key cell cycle functions, were correlated with viremia. The most profoundly upregulated biological functions related to cell cycle regulation, in particular, CDCA7 may drive aberrant cell division, promoted by overexpressed E2F family proteins. Also upregulated were DNA repair and replication, microtubule and spindle organization, and immune activation and response. The interferome of acute HIV was characterized by broad activation of interferon-stimulated genes with antiviral functions, most notably IFI27 and OTOF. BCL2 downregulation alongside upregulation of several apoptotic trigger genes and downstream effectors may contribute to cycle arrest and apoptosis. Transmembrane protein 155 (TMEM155) was consistently highly overexpressed during acute infection, with roles hitherto unknown. Discussion: Our study contributes to a better understanding of the mechanisms of early HIV-induced immune damage. These findings have the potential to lead to new earlier interventions that improve outcomes.

Correction to "An Iterative Approach Guides Discovery of the FabI Inhibitor Fabimycin, a Late-Stage Antibiotic Candidate with In Vivo Efficacy against Drug-Resistant Gram-Negative Infections".

[This corrects the article DOI: 10.1021/acscentsci.2c00598.].

An Iterative Approach Guides Discovery of the FabI Inhibitor Fabimycin, a Late-Stage Antibiotic Candidate with In Vivo Efficacy against Drug-Resistant Gram-Negative Infections.

Genomic studies and experiments with permeability-deficient strains have revealed a variety of biological targets that can be engaged to kill Gram-negative bacteria. However, the formidable outer membrane and promiscuous efflux pumps of these pathogens prevent many candidate antibiotics from reaching these targets. One such promising target is the enzyme FabI, which catalyzes the rate-determining step in bacterial fatty acid biosynthesis. Notably, FabI inhibitors have advanced to clinical trials for Staphylococcus aureus infections but not for infections caused by Gram-negative bacteria. Here, we synthesize a suite of FabI inhibitors whose structures fit permeation rules for Gram-negative bacteria and leverage activity against a challenging panel of Gram-negative clinical isolates as a filter for advancement. The compound to emerge, called fabimycin, has impressive activity against >200 clinical isolates of Escherichia coli, Klebsiella pneumoniae, and Acinetobacter baumannii, and does not kill commensal bacteria. X-ray structures of fabimycin in complex with FabI provide molecular insights into the inhibition. Fabimycin demonstrates activity in multiple mouse models of infection caused by Gram-negative bacteria, including a challenging urinary tract infection model. Fabimycin has translational promise, and its discovery provides additional evidence that antibiotics can be systematically modified to accumulate in Gram-negative bacteria and kill these problematic pathogens.

HIV infection in Eastern and Southern Africa: Highest burden, largest challenges, greatest potential.

BACKGROUND: The burden of HIV is especially concerning for Eastern and Southern Africa (ESA), as despite expansion of test-and-treat programmes, this region continues to experience significant challenges resulting from high rates of morbidity, mortality and new infections. Hard-won lessons from programmes on the ground in ESA should be shared. OBJECTIVES: This report summarises relevant evidence and regional experts' recommendations regarding challenges specific to ESA. METHOD: This commentary includes an in-depth review of relevant literature, progress against global goals and consensus opinion from experts. RESULTS: Recommendations include priorities for essential research (surveillance data collection, key and vulnerable population education and testing, in-country testing trials and evidence-based support services to improve retention in care) as well as research that can accelerate progress towards the prevention of new infections and achieving ambitious global goals in ESA. CONCLUSION: The elimination of HIV in ESA will require continued investment, commitment to evidence-based programmes and persistence. Local research is critical to ensuring that responses in ESA are targeted, efficient and evaluated.

Synthesis of Fusidic Acid Derivatives Yields a Potent Antibiotic with an Improved Resistance Profile.

Fusidic acid (FA) is a potent steroidal antibiotic that has been used in Europe for more than 60 years to treat a variety of infections caused by Gram-positive pathogens. Despite its clinical success, FA requires significantly elevated dosing (3 g on the first day, 1.2 g on subsequent days) to minimize resistance, as FA displays a high resistance frequency, and a large shift in minimum inhibitory concentration is observed for resistant bacteria. Despite efforts to improve on these aspects, all previously constructed derivatives of FA have worse antibacterial activity against Gram-positive bacteria than the parent natural product. Here, we report the creation of a novel FA analogue that has equivalent potency against clinical isolates of Staphylococcus aureus (S. aureus) and Enterococcus faecium (E. faecium) as well as an improved resistance profile in vitro when compared to FA. Importantly, this new compound displays efficacy against an FA-resistant strain of S. aureus in a soft-tissue murine infection model. This work delineates the structural features of FA necessary for potent antibiotic activity and demonstrates that the resistance profile can be improved for this scaffold and target.

Adapting Personal Therapies Using a Mobile Application for Tinnitus Rehabilitation: A Preliminary Study.

OBJECTIVES: To develop a smartphone application providing sound therapy and cognitive behavioral therapy (CBT) for treating tinnitus and performing a proof-of-concept pilot study evaluating its potential efficacy. METHODS: An interactive smartphone application available on iOS and Android platforms was developed, which provided an 8-week tinnitus-specific CBT and personalized and frequency-matched sound therapy. Included patients presented to our tertiary clinic between 2017 and 2018, while those waitlisted were regarded as controls. Three surveys were administrated: Tinnitus Handicap Inventory (THI), Generalized Anxiety Disorder 7-item (GAD-7), and Perceived Stress Scale (PSS). RESULTS: A total of 30 patients enrolled in this study consisting of 20 treatment and 10 control patients and mean age was 55.4 ± 11.6 years. Treatment and control patients had similar age, sex, and pre-enrolment GAD and PSS (all P > .05). Baseline THI scores were also similar between treatment and control cohorts (50.1 ± 21.9 vs 62.0 ± 20.7; P = .15). After 8 weeks, though changes in GAD and PSS scores were similar (P > .05), the treatment group reported a significantly greater improvement in THI scores (17.7 ± 15.8 vs 5.3 ± 10.5, P = .04). CONCLUSIONS: This pilot study demonstrated potentially promising efficacy of a smartphone-based CBT and sound therapy platform for treating tinnitus and encourages future randomized controlled trials on this treatment modality.

Gene Expression: the Key to Understanding HIV-1 Infection?

Gene expression profiling of the host response to HIV infection has promised to fill the gaps in our knowledge and provide new insights toward vaccine and cure. However, despite 20 years of research, the biggest questions remained unanswered. A literature review identified 62 studies examining gene expression dysregulation in samples from individuals living with HIV. Changes in gene expression were dependent on cell/tissue type, stage of infection, viremia, and treatment status. Some cell types, notably CD4+ T cells, exhibit upregulation of cell cycle, interferon-related, and apoptosis genes consistent with depletion. Others, including CD8+ T cells and natural killer cells, exhibit perturbed function in the absence of direct infection with HIV. Dysregulation is greatest during acute infection. Differences in study design and data reporting limit comparability of existing research and do not as yet provide a coherent overview of gene expression in HIV. This review outlines the extraordinarily complex host response to HIV and offers recommendations to realize the full potential of HIV host transcriptomics.

A Longitudinal Analysis Reveals Early Activation and Late Alterations in B Cells During Primary HIV Infection in Mozambican Adults.

Primary HIV infection (PHI) and subsequent chronic infection alter B-cell compartment. However, longitudinal analysis defining the dynamics of B-cell alterations are still limited. We longitudinally studied B-cell subsets in individuals followed for 1 year after PHI (n = 40). Treated and untreated chronic HIV infected (n = 56) and HIV-uninfected individuals (n = 58) were recruited as reference groups at the Manhiça District in Mozambique. B cells were analyzed by multicolor flow-cytometry. Anti-HIV humoral response and plasma cytokines were assessed by ELISA or Luminex-based technology. A generalized activation of B cells induced by HIV occurs early after infection and is characterized by increases in Activated and Tissue-like memory cells, decreases in IgM-IgD- (switched) and IgM-only B cells. These alterations remain mostly stable until chronic infection and are reverted in part by ART. In contrast, other parameters followed particular dynamics: PD-1 expression in memory cells decreases progressively during the first year of infection, Transitional B cells expand at month 3-4 after infection, and Marginal zone-like B cells show a late depletion. Plasmablasts expand 2 months after infection linked to plasma viral load and anti-p24 IgG3 responses. Most of well-defined changes induced by HIV in B-cell activation and memory subsets are readily observed after PHI, lasting until ART initiation. However, subsequent changes occur after sustained viral infection. These data indicate that HIV infection impacts B cells in several waves over time, and highlight that early treatment would result in beneficial effects on the B-cell compartment.

Implementation of permeation rules leads to a FabI inhibitor with activity against Gram-negative pathogens.

Gram-negative bacterial infections are a significant public health concern, and the lack of new drug classes for these pathogens is linked to the inability of most drug leads to accumulate inside Gram-negative bacteria1-7. Here, we report the development of a web application-eNTRyway-that predicts compound accumulation (in Escherichia coli) from its structure. In conjunction with structure-activity relationships and X-ray data, eNTRyway was utilized to re-design Debio-1452-a Gram-positive-only antibiotic8-into versions that accumulate in E. coli and possess antibacterial activity against high-priority Gram-negative pathogens. The lead compound Debio-1452-NH3 operates as an antibiotic via the same mechanism as Debio-1452, namely potent inhibition of the enoyl-acyl carrier protein reductase FabI, as validated by in vitro enzyme assays and the generation of bacterial isolates with spontaneous target mutations. Debio-1452-NH3 is well tolerated in vivo, reduces bacterial burden in mice and rescues mice from lethal infections with clinical isolates of Acinetobacter baumannii, Klebsiella pneumoniae and E. coli. This work provides tools for the facile discovery and development of high-accumulating compounds in E. coli, and a general blueprint for the conversion of Gram-positive-only compounds into broad-spectrum antibiotics.

Evaluation of a point-of-care ultrasound curriculum taught by medical students for physicians, nurses, and midwives in rural Indonesia.

PURPOSE: In the Indonesian health-care system, nurses and midwives often serve as the primary health-care providers due to physician shortages. Seeking to address the need for medical care in resource-limited environments, some have advocated for portable equipment in the hands of health-care providers. We hypothesized that medical students are able to effectively teach point-of-care ultrasound (POCUS) to physicians, nurses, and midwives in rural Indonesia. METHODS: We conducted a prospective, observational study using health-care practitioners from a clinic and accredited school for nursing and midwifery in Mojokerto, East Java, Indonesia. Enrolled practitioners took part in a 4-week POCUS course followed by postinstructional testing. RESULTS: A total of 55 health-care practitioners completed the course. This included 19 physicians, 13 nurses, and 19 midwives. Of the 55 clinicians, 43 (72%) passed the course and 12 (28%) failed. CONCLUSIONS: Physicians, nurses, and midwives in rural Indonesia showed significant acquisition of ultrasound (US) knowledge and skills following a 4-week US course. Following training, all three groups displayed skills in practical US use during a postcourse practical examination. This is one of the first studies to assess the efficacy of medical students teaching POCUS to midwives and nurses.

Efficacy of Multi-Modal Migraine Prophylaxis Therapy on Hyperacusis Patients.

OBJECTIVES: To evaluate the efficacy of a multi-modal migraine prophylaxis therapy for patients with hyperacusis. METHODS: In a prospective cohort, patients with hyperacusis were treated with a multi-modal step-wise migraine prophylactic regimen (nortriptyline, verapamil, topiramate, or a combination thereof) as well as lifestyle and dietary modifications. Pre- and post-treatment average loudness discomfort level (LDL), hyperacusis discomfort level measured by a visual analogue scale (VAS), and scores on the modified Khalfa questionnaire for severity of hyperacusis were compared. RESULTS: Twenty-two of the 25 patients (88%) reported subjective resolution of their symptoms following treatment. Post-treatment audiograms showed significant improvement in average LDL from 81.3 ± 3.2 dB to 86.4 ± 2.6 dB (P < .001), indicating increased sound tolerability. The VAS discomfort level also showed significant improvement from a pre-treatment average of 7.7 ± 1.1 to 3.7 ± 1.6 post-treatment (P < .001). There was also significant improvement in the average total score on modified Khalfa questionnaire (32.2 ± 3.6 vs 22.0 ± 5.7, P < .001). CONCLUSIONS: The majority of patients with hyperacusis demonstrated symptomatic improvement from migraine prophylaxis therapy, as indicated by self-reported and audiometric measures. Our findings indicate that, for some patients, hyperacusis may share a pathophysiologic basis with migraine disorder and may be successfully managed with multimodal migraine prophylaxis therapy.

IP-10 Levels as an Accurate Screening Tool to Detect Acute HIV Infection in Resource-Limited Settings.

Acute HIV infection (AHI) is the period prior to seroconversion characterized by high viral replication, hyper-transmission potential and commonly, non-specific febrile illness. AHI detection requires HIV-RNA viral load (VL) determination, which has very limited access in low-income countries due to restrictive costs and implementation constraints. We sought to identify a biomarker that could enable AHI diagnosis in scarce-resource settings, and to evaluate the feasibility of its implementation. HIV-seronegative adults presenting at the Manhiça District Hospital, Mozambique, with reported-fever were tested for VL. Plasma levels of 49 inflammatory biomarkers from AHI (n = 61) and non-HIV infected outpatients (n = 65) were determined by Luminex and ELISA. IP-10 demonstrated the best predictive power for AHI detection (AUC = 0.88 [95%CI 0.80-0.96]). A cut-off value of IP-10 ≥ 161.6 pg/mL provided a sensitivity of 95.5% (95%CI 85.5-99.5) and a specificity of 76.5% (95%CI 62.5-87.2). The implementation of an IP-10 screening test could avert from 21 to 84 new infections and save from US$176,609 to US$533,467 to the health system per 1,000 tested patients. We conclude that IP-10 is an accurate biomarker to screen febrile HIV-seronegative individuals for subsequent AHI diagnosis with VL. Such an algorithm is a cost-effective strategy to prevent disease progression and a substantial number of further HIV infections.

A Cytokine Pattern That Differentiates Preseroconversion From Postseroconversion Phases of Primary HIV Infection.

BACKGROUND: During acute HIV infection, HIV actively replicates but seroconversion has not yet occurred. Primary HIV infection (PHI) is characterized by a transient nonspecific febrile illness, a massive inflammatory response, and the progressive appearance of anti-HIV-specific antibodies. In this study, we have identified patterns of inflammatory biomarkers associated with the innate immunological reaction before completion of a full humoral response. METHODS: A symptom-based screening was used to identify acute HIV infection in the Manhiça District Hospital in Mozambique. Plasma levels of biomarkers were determined by Luminex and enzyme-linked immunosorbent assay. Anti-HIV antibodies were analyzed by flow cytometry and Western blot. Statistical analyses used random forest and logistic regression models. RESULTS: Of 3116 rapid test seronegative or indeterminate individuals, 85 (2.7%) had positive plasma HIV viral load and were enrolled as PHI, of which n = 45 (52.9%), n = 8 (9.4%), n = 12 (14.1%), and n = 20 (23.5%) were classified as Fiebig I-III, IV, V, and VI stages, respectively, by Western blot. Comparison of individuals at early (Fiebig I-IV) and late (Fiebig V-VI) immune stages identified significant differences in the expression level of plasma B-cell activating factor , monocyte chemotactic protein-1, sCD163, and monokine induced by interferon (IFN-γ). This cytokine signature classified patients in the preseroconversion phase with a sensitivity of 92.5% and a specificity of 81.2% CONCLUSIONS:: Identification of a cytokine signature specific for the preseroconversion stage of PHI may help to understand the earliest HIV pathogenic events and identify new potential targets for immunotherapy aimed at modulating the cytokine response to HIV infection.

Synthesis and biological evaluation of a water-soluble phosphate prodrug salt and structural analogues of KGP94, a lead inhibitor of cathepsin L.

The magnitude of expression of cathepsin L, often upregulated in the tumor microenvironment, correlates with the invasive and metastatic nature of certain tumors. Inhibition of cathepsin L represents an emerging strategy for the treatment of metastatic cancer. A potent, small-molecule inhibitor (referred to as KGP94) of cathepsin L, and new KGP94 analogues were synthesized. (3,5-Dibromophenyl)-(3-hydroxyphenyl) ketone thiosemicarbazone (22), with an IC50 value of 202nM, exhibited similar inhibitory activity against cathepsin L compared to KGP94 (IC50=189nM). Due to limited aqueous solubility of KGP94, a water-soluble phosphate salt (KGP420) was prepared in order to facilitate future in vivo studies. Enzymatic hydrolysis with alkaline phosphatase (ALP) demonstrated that the phosphate prodrug, KGP420, was readily converted to the parent compound, KGP94.

Dynamics of CD4 and CD8 T-Cell Subsets and Inflammatory Biomarkers during Early and Chronic HIV Infection in Mozambican Adults.

During primary HIV infection (PHI), there is a striking cascade response of inflammatory cytokines and many cells of the immune system show altered frequencies and signs of extensive activation. These changes have been shown to have a relevant role in predicting disease progression; however, the challenges of identifying PHI have resulted in a lack of critical information about the dynamics of early pathogenic events. We studied soluble inflammatory biomarkers and changes in T-cell subsets in individuals at PHI (n = 40), chronic HIV infection (CHI, n = 56), and HIV-uninfected (n = 58) recruited at the Manhiça District Hospital in Mozambique. Plasma levels of 49 biomarkers were determined by Luminex and ELISA. T-cell immunophenotyping was performed by multicolor flow cytometry. Plasma HIV viremia, CD4, and CD8 T cell counts underwent rapid stabilization after PHI. However, several immunological parameters, including Th1-Th17 CD4 T cells and activation or exhaustion of CD8 T cells continued decreasing until more than 9 months postinfection. Importantly, no sign of immunosenescence was observed over the first year of HIV infection. Levels of IP-10, MCP-1, BAFF, sCD14, tumor necrosis factor receptor-2, and TRAIL were significantly overexpressed at the first month of infection and underwent a prompt decrease in the subsequent months while, MIG and CD27 levels began to increase 1 month after infection and remained overexpressed for almost 1 year postinfection. Early levels of soluble biomarkers were significantly associated with subsequently exhausted CD4 T-cells or with CD8 T-cell activation. Despite rapid immune control of virus replication, the stabilization of the T-cell subsets occurs months after viremia and CD4 count plateau, suggesting persistent immune dysfunction and highlighting the potential benefit of early treatment initiation that could limit immunological damage.

Synthesis and biochemical evaluation of benzoylbenzophenone thiosemicarbazone analogues as potent and selective inhibitors of cathepsin L.

Upregulation of cathepsin L in a variety of tumors and its ability to promote cancer cell invasion and migration through degradation of the extracellular matrix suggest that cathepsin L is a promising biological target for the development of anti-metastatic agents. Based on encouraging results from studies on benzophenone thiosemicarbazone cathepsin inhibitors, a series of fourteen benzoylbenzophenone thiosemicarbazone analogues were designed, synthesized, and evaluated for their inhibitory activity against cathepsins L and B. Thiosemicarbazone inhibitors 3-benzoylbenzophenone thiosemicarbazone 1, 1,3-bis(4-fluorobenzoyl)benzene thiosemicarbazone 8, and 1,3-bis(2-fluorobenzoyl)-5-bromobenzene thiosemicarbazone 32 displayed the greatest potency against cathepsin L with low IC50 values of 9.9 nM, 14.4 nM, and 8.1 nM, respectively. The benzoylbenzophenone thiosemicarbazone analogues evaluated were selective in their inhibition of cathepsin L compared to cathepsin B. Thiosemicarbazone analogue 32 inhibited invasion through Matrigel of MDA-MB-231 breast cancer cells by 70% at 10 µM. Thiosemicarbazone analogue 8 significantly inhibited the invasive potential of PC-3ML prostate cancer cells by 92% at 5 µM. The most active cathepsin L inhibitors from this benzoylbenzophenone thiosemicarbazone series (1, 8, and 32) displayed low cytotoxicity toward normal primary cells [in this case human umbilical vein endothelial cells (HUVECs)]. In an initial in vivo study, 3-benzoylbenzophenone thiosemicarbazone (1) was well-tolerated in a CDF1 mouse model bearing an implanted C3H mammary carcinoma, and showed efficacy in tumor growth delay. Low cytotoxicity, inhibition of cell invasion, and in vivo tolerability are desirable characteristics for anti-metastatic agents functioning through an inhibition of cathepsin L. Active members of this structurally diverse group of benzoylbenzophenone thiosemicarbazone cathepsin L inhibitors show promise as potential anti-metastatic, pre-clinical drug candidates.

Rapid HIV progression during acute HIV-1 subtype C infection in a Mozambican patient with atypical seroconversion.

Acute human immunodeficiency virus (HIV) infection (AHI) refers to the period between viral transmission and development of an adaptive immune response to HIV antigens (seroconversion) usually lasting 6-8 weeks. Rare cases have been described in which HIV-infected patients fail to seroconvert and instead, develop rapid HIV-mediated clinical decline. We report the case of a Mozambican woman with AHI and malaria coinfection who showed atypical seroconversion and experienced rapid deterioration and death within 14 weeks of diagnosis with AHI. Atypical seroconversion may be associated with rapid progression. Fourth generation rapid tests could lead to earlier identification and intervention for this vulnerable subgroup.

Initial evaluation of the antitumour activity of KGP94, a functionalized benzophenone thiosemicarbazone inhibitor of cathepsin L.

Kinetic analysis of the mode of inhibition of cathepsin L by KGP94, a lead compound from a privileged library of functionalized benzophenone thiosemicarbazone derivatives, demonstrated that it is a time-dependent, reversible, and competitive inhibitor of the enzyme. These results are consistent with the formation of a transient covalent bond, and are supported by molecular modeling that places the thiocarbonyl of the inhibitor in proximity to the thiolate moiety of the enzyme active site Cys25. KGP94 significantly decreased the activity of cathepsin L toward human type I collagen, and impeded both migration and invasion of MDA-MB-231 human breast cancer cells. Growth retardation was achieved in vivo against both recently implanted and established tumours using a C3H mouse mammary carcinoma model.