Chimeric hepatitis B surface antigen virus-like particles expressing the strep A epitope p*17 elicit a humoral immune response in mice.

To optimize immunogenicity, bacterial epitopes in putative vaccine constructs can be presented to immune cells as multiple repeated structures on a defined nanoparticle. Virus-like particles (VLPs) are viral capsid proteins that self-assemble to form compact and highly ordered nanoparticles that are within the optimal size range for uptake by dendritic cells. VLPs mimic the live virus in size and form but contain no viral genetic material, are therefore noninfective and are the basis of safe and effective vaccines against hepatitis B virus (HBV) and human papillomavirus (HPV). Due to their particulate nature, molecular stability, and expression of high density and repetitive antigen displays, recombinant cell culture-derived VLPs are ideal platforms for the delivery of small molecules, including bacterial epitopes. We developed a putative vaccine by expressing a minimal epitope from the bacterium Streptococcus pyogenes (Strep A) on the surface of a recombinant VLP comprising multiple copies of HBV small envelope protein (HBsAg-S). Strep A is responsible for a wide spectrum of human infections and postinfectious diseases that disproportionately affect children and young adults living in resource-poor communities. No vaccine is currently available to offer sufficiently broad protection from the numerous and diverse strains of Strep A endemic in these at-risk populations. The Strep A antigen targeted by our vaccine construct is p*17, a cryptic epitope from a highly conserved region of the Strep A M-protein with demonstrated enhanced immunogenicity and broad protective potential against Strep A. To ensure surface expression and optimal immunogenicity, we expressed p*17 within the immunodominant "a" determinant of HBsAg-S. The recombinant VLPs (VLP-p*17) expressed in HEK293T cells spontaneously formed 22 nm particles and induced the production of high titers of p*17-specific IgG in BALB/c mice immunized with three 0.5 µg doses of VLP-p*17 formulated with adjuvant.

Detection, characterization, and persistence of Campylobacter hepaticus, the cause of spotty liver disease in layer hens.

A Campylobacter species was first described as the etiological agent of Spotty Liver Disease (SLD) in 2015 and subsequently named as Campylobacter hepaticus in 2016. The bacterium predominantly affects barn and/or free-range hens at peak lay, is fastidious and difficult to isolate, which has impeded elucidation of its sources, means of persistence and transmission. Ten farms from South-Eastern Australia, of which 7 were free range entities participated in the study. A total of 1,404 specimens from layers and 201 from environmental sources, were examined for the presence of C. hepaticus. In this study, our principal findings included the continuing detection of C. hepaticus infection in a flock following an outbreak, indicating a possible transition of infected hens to asymptomatic carriers, that was also characterized by no further occurrence of SLD in the flock. We also report that the first outbreaks of SLD on newly commissioned free-range farms affected layers ranging from 23 to 74 wk of age, while subsequent outbreaks in replacement flocks on these farms occurred during the more conventional peak lay period (23-32 wk of age). Finally, we report that in the on-farm environment, C. hepaticus DNA was detected in layer feces, inert elements such as stormwater, mud, soil, as well as in fauna such as flies, red mites, Darkling beetles, and rats. While in off-farm locations, the bacterium was detected in feces from a variety of wild birds and a canine.

Integration of immunoinformatics and cheminformatics to design and evaluate a multitope vaccine against Klebsiella pneumoniae and Pseudomonas aeruginosa coinfection.

Introduction: Klebsiella pneumoniae (K. pneumoniae) and Pseudomonas aeruginosa (P. aeruginosa) are the most common Gram-negative bacteria associated with pneumonia and coinfecting the same patient. Despite their high virulence, there is no effective vaccine against them. Methods: In the current study, the screening of several proteins from both pathogens highlighted FepA and OmpK35 for K. pneumonia in addition to HasR and OprF from P. aeruginosa as promising candidates for epitope mapping. Those four proteins were linked to form a multitope vaccine, that was formulated with a suitable adjuvant, and PADRE peptides to finalize the multitope vaccine construct. The final vaccine's physicochemical features, antigenicity, toxicity, allergenicity, and solubility were evaluated for use in humans. Results: The output of the computational analysis revealed that the designed multitope construct has passed these assessments with satisfactory scores where, as the last stage, we performed a molecular docking study between the potential vaccine construct and K. pneumonia associated immune receptors, TLR4 and TLR2, showing affinitive to both targets with preferentiality for the TLR4 receptor protein. Validation of the docking studies has proceeded through molecular dynamics simulation, which estimated a strong binding and supported the nomination of the designed vaccine as a putative solution for K. pneumoniae and P. aeruginosa coinfection. Here, we describe the approach for the design and assessment of our potential vaccine.

Study of the role of microRNAs 16 and 135a in patients with lifelong premature ejaculation receiving fluoxetine daily for 3 months: A prospective case control study.

We aimed to determine the level of miRNAs 16 and 135a in lifelong premature ejaculation (LPE) patients versus controls. Moreover, we evaluated the potential interplay between the studied miRNAs and fluoxetine in these patients after utilizing fluoxetine daily for 3 months. The study involved 60 consecutive LPE patients and 20 healthy age matched individuals as controls. The median miRNA16 was significantly higher in the controls (1.02) compared to the patients (0.31) (p < 0.001). Moreover, the median miRNA-135a was significantly higher in the controls compared to the patients 1.02 and 0.35, p < 0.001, respectively. In addition, the median pre-treatment miRNA16 in the responders was 0.29 that significantly increased to 0.66 (p < 0.001). The median pre-treatment miRNA-135a in the responders was 0.27 that significantly increased to 0.65 (p < 0.001). Furthermore, considering EXP(ß) for the odds ratio evaluation, with a 95% degree of confidence, a 1 fold increase in pre-treatment miRNA 135a fold change decreases the odds for being responsive to SSRI by 0.028. Meanwhile, there was non-significant association between fluoxetine responsiveness and age, pre-treatment miRNA 16, pre-treatment PEDT and pre-treatment IELT. The current study had shown that a lower pre-treatment miRNA 135a was significantly associated with response to fluoxetine.

Evaluation of the sensitivity of different doses of vasoactive drugs in diagnosing erectile dysfunction in impotent patients: a prospective case-control study.

INTRODUCTION: Erectile dysfunction (ED) is one of the most common sexual disorders worldwide affecting about 30 million men in the United States, and an estimated 100 million men worldwide. Penile duplex doppler ultrasound (PDDU) is performed using an intracavernosal injection (ICI) of a vasoactive agent to demonstrate both arterial insufficiency and veno-occlusive dysfunction. This article aims to evaluate the sensitivity of different doses of different vasoactive agents used to diagnose ED in impotent patients. MATERIAL AND METHODS: This study recruited 90 subjects with ED and 100 healthy subjects as controls. All of the subjects were assessed using the International Index of Erectile Function score (IIEF-5) while degree of erection was assessed by the Erection Hardness Score (EHS). Two penile duplex tests were done for each candidate two weeks apart. RESULTS: None of the sample population achieved a normal clinical response (EHS >2) to 10 ug PGE1. In contrast, 60 controls (60%) had a normal response (EHS >2) to 10 ug PGE1. This difference in response between the sample and control populations to 10 ug PGE1 was of high statistical significance 11 (p <0.001). In contrast, 54 (60%) out of the 90 cases had normal clinical response (EHS >2) to 0.25cc Trimix (everywhere). Interestingly, 96 controls (96%) demonstrated normal response (EHS >2) to 0.25cc Trimix. This difference in response between the sample and control populations to 0.25 cc Trimix was also of high statistical significance (p <0.001). CONCLUSIONS: Our study demonstrated a statistically significant association between the response to Trimix over PGE1 and peak systolic velocity (PSV) and end diastolic velocity (EDV). Thus, we conclude that 0.25 cc Trimix is more sensitive than 20 ug PGE1 in diagnosing ED for impotent patients and also provides a more potent response.

Rheumatic heart disease: A review of the current status of global research activity.

Rheumatic heart disease (RHD) is a serious and long-term consequence of acute rheumatic fever (ARF), an autoimmune sequela of a mucosal infection by Streptococcus pyogenes (Group A Streptococcus, Strep A). The pathogenesis of ARF and RHD is complex and not fully understood but involves host and bacterial factors, molecular mimicry, and aberrant host innate and adaptive immune responses that result in loss of self-tolerance and subsequent cross-reactivity with host tissues. RHD is entirely preventable yet claims an estimated 320 000 lives annually. The major burden of disease is carried by developing nations and Indigenous populations within developed nations, including Australia. This review will focus on the epidemiology, pathogenesis and treatment of ARF and RHD in Australia, where: streptococcal pyoderma, rather than streptococcal pharyngitis, and Group C and Group G Streptococcus, have been implicated as antecedents to ARF; the rates of RHD in remote Indigenous communities are persistently among the highest in the world; government register-based programs coordinate disease screening and delivery of prophylaxis with variable success; and researchers are making significant progress in the development of a broad-spectrum vaccine against Strep A.

The development of ghost vaccines trials.

INTRODUCTION: Bacterial ghosts are intact bacterial cell envelopes that are emptied of their content by gentle biological or chemical poring methods. Ghost techniques increase the safety of the killed vaccines, while maintaining their antigenicity due to mild preparation procedures. Moreover, ghost-platforms may express and/or carry several antigens or plasmid-DNA encoding for protein epitopes. AREAS COVERED: In this review, the development in ghost-vaccine production over the last 30 years is classified and discussed. The different applications of ghost-vaccines, how they trigger the immune system, their advantages and limitations are displayed. The phage-mediated lysis, molecular manipulation of the lysis-genes, and the biotechnological production of ghosts are described. The trials are classified according to the pattern of lysis and to the type of bacteria. Further subdivision includes chronological ordered application of the ghost as alternative-killed vaccine, recombinant antigen platform, plasmid DNA carrier, adjuvants, and dendritic cell inducer. Particular trials for specific pathogens or from distinct research schools are gathered. EXPERT OPINION: Ghosts are highly qualified to act as immune-presenting platforms that express and/or carry several recombinant and DNA vaccines, as well as, being efficient alternative-killed vaccines. The coming years will show more molecular advances to develop ghost-production and to express more antigens.

Effect of Time of Day of Infection on Chlamydia Infectivity and Pathogenesis.

Genital chlamydia infection in women causes complications such as pelvic inflammatory disease and tubal factor infertility, but it is unclear why some women are more susceptible than others. Possible factors, such as time of day of chlamydia infection on chlamydial pathogenesis has not been determined. We hypothesised that infections during the day, will cause increased complications compared to infections at night. Mice placed under normal 12:12 light: dark (LD) cycle were infected intravaginally with Chlamydia muridarum either at zeitgeber time 3, ZT3 and ZT15. Infectivity was monitored by periodic vaginal swabs and chlamydiae isolation. Blood and vaginal washes were collected for host immunologic response assessments. The reproductive tracts of the mice were examined histopathologically, and fertility was determined by embryo enumeration after mating. Mice infected at ZT3 shed significantly more C. muridarum than mice infected at ZT15. This correlated with the increased genital tract pathology observed in mice infected at ZT3. Mice infected at ZT3 were less fertile than mice infected at ZT15. The results suggest that the time of day of infection influences chlamydial pathogenesis, it indicates a possible association between complications from chlamydia infection and host circadian clock, which may lead to a better understanding of chlamydial pathogenesis.

Correlation between Fatty Acid Profile and Anti-Inflammatory Activity in Common Australian Seafood by-Products.

Marine organisms are a rich source of biologically active lipids with anti-inflammatory activities. These lipids may be enriched in visceral organs that are waste products from common seafood. Gas chromatography-mass spectrometry and fatty acid methyl ester (FAME) analyses were performed to compare the fatty acid compositions of lipid extracts from some common seafood organisms, including octopus (Octopus tetricus), squid (Sepioteuthis australis), Australian sardine (Sardinops sagax), salmon (Salmo salar) and school prawns (Penaeus plebejus). The lipid extracts were tested for anti-inflammatory activity by assessing their inhibition of nitric oxide (NO) and tumor necrosis factor alpha (TNFα) production in lipopolysaccharide (LPS)-stimulated RAW 264.7 mouse cells. The lipid extract from both the flesh and waste tissue all contained high amounts of polyunsaturated fatty acids (PUFAs) and significantly inhibited NO and TNFα production. Lipid extracts from the cephalopod mollusks S. australis and O. tetricus demonstrated the highest total PUFA content, the highest level of omega 3 (ω-3) PUFAs, and the highest anti-inflammatory activity. However, multivariate analysis indicates the complex mixture of saturated, monounsaturated, and polyunsaturated fatty acids may all influence the anti-inflammatory activity of marine lipid extracts. This study confirms that discarded parts of commonly consumed seafood species provide promising sources for the development of new potential anti-inflammatory nutraceuticals.

The Enhancement of the Pasteurella's Bacterin by Propolis Extracts.

BACKGROUND: Pasteurella multocida continues to pose a danger to prone farm and wild animals all over the world. Chemotherapeutic treatments are progressively losing their effectiveness, last for long time, and cost a lot of money, as well as being toxic to human consumers. Therefore, clearing the way for immunization as a big-wheel alternative against the economic grain. Yet, the vaccines available in the market do not confer the necessary protection against the pathogen. The integration of the well adjuvanted killed vaccine with the attenuated vaccines proved to offer an effective protection to the host animals. However, the bare use of the killed bacterin to provide protection from the possible harm of the live attenuated vaccine was doubtful. METHODS: In the present study, propolis extracts were used to ameliorate the immunogenicity of the Pasteurella bacterin. The cellular and humoral activities were assessed for the different bacterin formulations. RESULTS: Propolis extracts adjuvants proved to broaden and extend the IgG potency, as well as to induce a unique mucosal protection against the bacterium. Simultaneously it offered an anti-inflammatory effect that increased the tolerability to the bacterin. While the cellular activity was relatively reduced with propolis extracts. CONCLUSION: These results confirm the effectiveness of the formulation of the bacterin with propolis to offer a potent homologous primary protection to the animals against the long-life use of the attenuated Pasteurella vaccines.

Review of anti-inflammatory, immune-modulatory and wound healing properties of molluscs.

ETHNOPHARMACOLOGICAL RELEVANCE: This review focuses on traditional and contemporary anti-inflammatory uses of mollusc-derived products summarising all the in vitro, in vivo and human clinical trials that have tested the anti-inflammatory activity of molluscan natural products. Inflammatory conditions, burns and wounds have been an ongoing concern for human health since the early era of civilisation. Many texts from ancient medicine have recorded the symptoms, signs and treatments for these conditions. Natural treatments are well-documented in traditional European medicine, Traditional Chinese Medicine (TCM), Siddha and ancient Mediterranean and African traditional medicine and include a surprisingly large number of molluscan species. MATERIALS AND METHODS: An extensive review of the Materia Medica and scientific literature was undertaken using key word searches for "mollusc" and "anti-inflammatory" or "immunomodulatory" or "wound healing". RESULTS: Molluscs have been used in ethnomedicine by many traditional cultures to treat different aspects of inflammatory conditions. We found 104 different anti-inflammatory preparations from a variety of molluscan species, of which 70 were from the well-documented Traditional Chinese Medicine (TCM). This traditional use of molluscs has driven the testing for inflammatory activity in extracts from some species in the phylum Mollusca, with 20 in vitro studies, 40 in vivo animal studies and 14 human clinical trials performed to substantiate the anti-inflammatory and wound healing activity of molluscs. Some of these studies have led to the approval of mollusc-derived products to be used as over-the-counter (OTC) nutraceuticals, like Lyprinol® and Biolane™ from the New Zealand green lipped mussel Perna canaliculus. CONCLUSION: Natural products provide important leads for the development of pharmaceuticals, including anti-inflammatory agents. Only a small proportion of the molluscan traditional medicines have been tested to confirm their anti-inflammatory activity and most screening studies have tested crude extracts from molluscs without any chemical characterisation. This highlights the need for further research to strategically identify the anti-inflammatory compounds in molluscan medicines to provide leads for novel anti-inflammatory drugs in the future.

Brominated indoles from a marine mollusc inhibit inflammation in a murine model of acute lung injury.

New drug leads for the treatment of inflammation are urgently needed. Marine molluscs are widely used as traditional medicines for the treatment of inflammation. Here we report the positive effects of a hypobranchial gland (HBG) extract and the dominant bioactive compound 6-bromoisatin from the Muricidae mollusc Dicathais orbita, for reducing lipopolysaccharide (LPS) induced acute lung inflammation in a mouse model. Both 6-bromoisatin and the HBG extract suppressed the inflammatory response in mice that were pre-treated by oral gavage at 48, 24 and 1 h prior to LPS infusion. The inflammatory antagonists were tested at concentrations of 0.5 mg/g and 0.1 mg/g HBG extract and 0.1 mg/g and 0.05 mg/g 6-bromoisatin in carrier oil and all treatments reduced inflammation as indicated by a significant suppression of inflammatory markers present in bronchoalveolar lavage fluid (BALF), in comparison to LPS induced positive control mice administered the carrier oil alone (p < 0.0001). Tumour necrosis factor-alpha (TNFα) and interleukin-1 beta (IL-1ß) levels, in addition to total protein concentration were all significantly reduced in BALF from mice treated with the extract or 6-bromoisatin. Furthermore, all treatment groups showed significant reductions in neutrophil sequestration and preservation of the lung tissue architecture compared to the positive control (p < 0.0001). The combined results from this study and our previous in vitro studies indicate that 6-bromoisatin in the HGB extracts inhibit the activation of inflammatory signalling pathway. The results from this study further confirm that the HBG extract from Muricidae molluscs and 6-bromoisatin are bioavailable and effective in vivo, thus have potential for development as natural therapeutic agents for inflammation.

Anti-Inflammatory Activity and Structure-Activity Relationships of Brominated Indoles from a Marine Mollusc.

Marine molluscs are rich in biologically active natural products that provide new potential sources of anti-inflammatory agents. Here we used bioassay guided fractionation of extracts from the muricid Dicathais orbita to identify brominated indoles with anti-inflammatory activity, based on the inhibition of nitric oxide (NO) and tumour necrosis factor α (TNFα) in lipopolysaccharide (LPS) stimulated RAW264.7 macrophages and prostaglandin E2 (PGE2) in calcium ionophore-stimulated 3T3 ccl-92 fibroblasts. Muricid brominated indoles were then compared to a range of synthetic indoles to determine structure-activity relationships. Both hypobranchial gland and egg extracts inhibited the production of NO significantly with IC50 of 30.8 and 40 µg/mL, respectively. The hypobranchial gland extract also inhibited the production of TNFα and PGE2 with IC50 of 43.03 µg/mL and 34.24 µg/mL, respectively. The purified mono-brominated indole and isatin compounds showed significant inhibitory activity against NO, TNFα, and PGE2, and were more active than dimer indoles and non-brominated isatin. The position of the bromine atom on the isatin benzene ring significantly affected the activity, with 5Br > 6Br > 7Br. The mode of action for the active hypobranchial gland extract, 6-bromoindole, and 6-bromoisatin was further tested by the assessment of the translocation of nuclear factor kappa B (NFκB) in LPS-stimulated RAW264.7 mouse macrophage. The extract (40 µg/mL) significantly inhibited the translocation of NFκB in the LPS-stimulated RAW264.7 macrophages by 48.2%, whereas 40 µg/mL of 6-bromoindole and 6-bromoistain caused a 60.7% and 63.7% reduction in NFκB, respectively. These results identify simple brominated indoles as useful anti-inflammatory drug leads and support the development of extracts from the Australian muricid D. orbita, as a new potential natural remedy for the treatment of inflammation.

The detection of antigenic determinants of Acinetobacter baumannii.

BACKGROUND: Acinetobacter baumannii continues to pose a threat to burdened patients in ICUs all around the world. Lately, infection control techniques are not sufficient to curb A. baumannii's progression and chemotherapeutics are losing their potency against it. Thus, immunization became a key player in providing an ideal solution to the dilemma. None of the vaccines under investigation have reached the market and the search for a tailored vaccine remains a challenge. The notion of unravelling the bacterial antigens to design a novel epitope-based vaccine proved its merits. METHODS: In this work, the propitious polysaccharide and protein antigenic determinants of A. baumannii were mapped by mimicking the infection. The immune response was evaluated by western blot, ELISA, and cellular proliferation assay techniques. RESULTS: The screening showed that OMPs induced the most eminent sustained IgG response. In addition, OMP gave the highest cellular proliferation and a fold increase in ELISA that reached up to 10-fold by week 6. Whilst, the LPS gave a rapid IgM response, that reached 5-fold and the response was visible from week 1 in the western blot. The OMPs had a more pronounced effect in eliciting a cellular immune response. CONCLUSION: The results elaborated the valuable role of using pure OMPs and detoxified LPS together; as a major cornerstone in designing an ideal vaccine against A. baumannii.

Development of immunization trials against Pasteurella multocida.

Pasteurellosis is one of the most important respiratory diseases facing economically valuable farm animals such as poultry, rabbit, cattle, goats and pigs. It causes severe economic loss due to its symptoms that range from primary local infection to fatal septicemia. Pasteurella multocida is the responsible pathogen for this contagious disease. Chemotherapeutic treatment of Pasteurella is expensive, lengthy, and ineffective due to the increasing antibiotics resistance of the bacterium, as well as its toxicity to human consumers. Though, biosecurity measures played a role in diminishing the spread of the pathogen, the immunization methods were always the most potent preventive measures. Since the early 1950s, several trials for constructing and formulating effective vaccines were followed. This up-to-date review classifies and documents such trials. A section is devoted to discussing each group benefits and defects.

The use of propolis as vaccine's adjuvant.

The success of many vaccines relies on their association with selected adjuvants in order to increase their immunogenicity and ensure long-term protection. All available adjuvants have adverse effects due to their toxicity and reactogenicity. Pre-clinical in vivo investigations can identify new natural products for further applications. Several studies have confirmed the different medicinal benefits of propolis. However the studies that addressed its use as a potent, safe, vaccine adjuvant were limited to specific countries and languages, primarily Chinese. Those studies introduced the use of different extracts and formulations of propolis as adjuvants for bacterial, viral, and parasitic vaccines. This comprehensive up-to-date review categorizes, documents, and discusses those trials in a clear chronological manner.

Development of a new trend conjugate vaccine for the prevention of Klebsiella pneumoniae.

Klebsiella pneumoniae is a major cause of nosocomial pneumonia, septicemia and urinary tract infections, especially in newborns, blood cancer patients, and other immunocompromised candidates. The control of K. pneumoniae is a complicated issue due to its tight pathogenesis. Immuno-prophylactic preparations, especially those directed toward the bacterium O-antigen, showed to be the most successful way to prevent the infection incidence. However, all previously proposed preparations were either of limited spectrum or non-maternal, and hence not targeting the main Klebsiella patients. Moreover, all preparations were directed only to prevent the respiratory diseases due to that pathogen. This article addresses the development of a method originally used to purify the non-capsular bacterial-endotoxins, as a new and easy method for vaccine production against K. pneumoniae. The application of this method was preceded by a biotechnological control of capsular polysaccharide production in K. pneumoniae. The new produced natural conjugate between the bacterial O-antigen and its outer membrane proteins was evaluated by physicochemical and immunological methods to investigate its purity, integrity, safety and immunogenicity. It showed to be pure, stable, safe for use, and able to elicit a protective immunoglobulin titer against different Klebsiella infections. This immune-response proved to be transferable to the offspring of the vaccinated experimental rabbits via placenta.

Development of immunization trials against Klebsiella pneumoniae.

Klebsiella pneumoniae is the most common cause of nosocomial respiratory tract and premature intensive care infections, and the second most frequent cause of Gram-negative bacteraemia and urinary tract infections. Drug resistant isolates remain an important hospital-acquired bacterial pathogen, add significantly to hospital stays, and are especially problematic in high impact medical areas such as intensive care units. Many investigations worldwide proved the increasing resistance of such pathogen, resulting in an average rate of 1.63 outbreak every year. A variety of preventive measures were applied to reduce such incidences. Immunotherapy and passive immunization researches as well found their way to the treatment of Klebsiella. During the last 40 years, many trials for constructing effective vaccines were followed. This up-to-date review classifies such trials and documents them in a progressive way. A following comment discusses each group benefits and defects.