Multivalent poultry vaccine development using Protein Glycan Coupling Technology

Abstract Background Poultry is the world's most popular animal-based food and global production has tripled in the past 20 years alone. Low-cost vaccines that can be combined to protect poultry against multiple infections are a current global imperative. Glycoconjugate vaccines, which consist of an immunogenic protein covalently coupled to glycan antigens of the targeted pathogen, have a proven track record in human vaccinology, but have yet to be used for livestock due to prohibitively high manufacturing costs. To overcome this, we use Protein Glycan Coupling Technology (PGCT), which enables the production of glycoconjugates in bacterial cells at considerably reduced costs, to generate a candidate glycan-based live vaccine intended to simultaneously protect against Campylobacter jejuni, avian pathogenic Escherichia coli (APEC) and Clostridium perfringens. Campylobacter is the most common cause of food poisoning, whereas colibacillosis and necrotic enteritis are widespread and devastating infectious diseases in poultry. Results We demonstrate the functional transfer of C. jejuni protein glycosylation (pgl) locus into the genome of APEC χ7122 serotype O78:H9. The integration caused mild attenuation of the χ7122 strain following oral inoculation of chickens without impairing its ability to colonise the respiratory tract. We exploit the χ7122 pgl integrant as bacterial vectors delivering a glycoprotein decorated with the C. jejuni heptasaccharide glycan antigen. To this end we engineered χ7122 pgl to express glycosylated NetB toxoid from C. perfringens and tested its ability to reduce caecal colonisation of chickens by C. jejuni and protect against intra-air sac challenge with the homologous APEC strain. Conclusions We generated a candidate glycan-based multivalent live vaccine with the potential to induce protection against key avian and zoonotic pathogens (C. jejuni, APEC, C. perfringens). The live vaccine failed to significantly reduce Campylobacter colonisation under the conditions tested but was protective against homologous APEC challenge. Nevertheless, we present a strategy towards the production of low-cost “live-attenuated multivalent vaccine factories” with the ability to express glycoconjugates in poultry.

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An atypical lipoteichoic acid from Clostridium perfringens elicits a broadly cross-reactive and protective immune response

Journal of Biological Chemistry ◽

10.1074/jbc.ra119.009978 ◽

2020 ◽

Vol 295 (28) ◽

pp. 9513-9530 ◽

Cited By ~ 1

Author(s):

Cory Q. Wenzel ◽

Dominic C. Mills ◽

Justyna M. Dobruchowska ◽

Jiri Vlach ◽

Harald Nothaft ◽

...

Keyword(s):

Clostridium Perfringens ◽

Vaccine Development ◽

Food Poisoning ◽

Laboratory Strain ◽

Lipoteichoic Acid ◽

2D Nmr ◽

Effective Vaccine ◽

Necrotic Enteritis ◽

Bacterial Killing ◽

Whole Cells

Clostridium perfringens is a leading cause of food-poisoning and causes avian necrotic enteritis, posing a significant problem to both the poultry industry and human health. No effective vaccine against C. perfringens is currently available. Using an antiserum screen of mutants generated from a C. perfringens transposon-mutant library, here we identified an immunoreactive antigen that was lost in a putative glycosyltransferase mutant, suggesting that this antigen is likely a glycoconjugate. Following injection of formalin-fixed whole cells of C. perfringens HN13 (a laboratory strain) and JGS4143 (chicken isolate) intramuscularly into chickens, the HN13-derived antiserum was cross-reactive in immunoblots with all tested 32 field isolates, whereas only 5 of 32 isolates were recognized by JGS4143-derived antiserum. The immunoreactive antigens from both HN13 and JGS4143 were isolated, and structural analysis by MALDI-TOF-MS, GC-MS, and 2D NMR revealed that both were atypical lipoteichoic acids (LTAs) with poly-(β1→4)-ManNAc backbones substituted with phosphoethanolamine. However, although the ManNAc residues in JGS4143 LTA were phosphoethanolamine-modified, a few of these residues were instead modified with phosphoglycerol in the HN13 LTA. The JGS4143 LTA also had a terminal ribose and ManNAc instead of ManN in the core region, suggesting that these differences may contribute to the broadly cross-reactive response elicited by HN13. In a passive-protection chicken experiment, oral challenge with C. perfringens JGS4143 lead to 22% survival, whereas co-gavage with JGS4143 and α-HN13 antiserum resulted in 89% survival. This serum also induced bacterial killing in opsonophagocytosis assays, suggesting that HN13 LTA is an attractive target for future vaccine-development studies.

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Platforms for Production of Protein-Based Vaccines: From Classical to Next-Generation Strategies

Biomolecules ◽

10.3390/biom11081072 ◽

2021 ◽

Vol 11 (8) ◽

pp. 1072

Author(s):

Raquel Cid ◽

Jorge Bolívar

Keyword(s):

Infectious Diseases ◽

Mammalian Cells ◽

Vaccine Development ◽

Low Cost ◽

Cost Effective ◽

Insect Larvae ◽

Subunit Vaccines ◽

Effective Strategies ◽

Alternative Systems ◽

Inactivated Vaccines

To date, vaccination has become one of the most effective strategies to control and reduce infectious diseases, preventing millions of deaths worldwide. The earliest vaccines were developed as live-attenuated or inactivated pathogens, and, although they still represent the most extended human vaccine types, they also face some issues, such as the potential to revert to a pathogenic form of live-attenuated formulations or the weaker immune response associated with inactivated vaccines. Advances in genetic engineering have enabled improvements in vaccine design and strategies, such as recombinant subunit vaccines, have emerged, expanding the number of diseases that can be prevented. Moreover, antigen display systems such as VLPs or those designed by nanotechnology have improved the efficacy of subunit vaccines. Platforms for the production of recombinant vaccines have also evolved from the first hosts, Escherichia coli and Saccharomyces cerevisiae, to insect or mammalian cells. Traditional bacterial and yeast systems have been improved by engineering and new systems based on plants or insect larvae have emerged as alternative, low-cost platforms. Vaccine development is still time-consuming and costly, and alternative systems that can offer cost-effective and faster processes are demanding to address infectious diseases that still do not have a treatment and to face possible future pandemics.

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Arthritis and Foodborne Bacteria

Journal of Food Protection ◽

10.4315/0362-028x-57.10.935 ◽

1994 ◽

Vol 57 (10) ◽

pp. 935-941 ◽

Cited By ~ 11

Author(s):

JAMES L. SMITH

Keyword(s):

T Cells ◽

Ankylosing Spondylitis ◽

Reactive Arthritis ◽

Molecular Mimicry ◽

Food Poisoning ◽

Cell Subset ◽

Bacterial Cells ◽

Hla B27 ◽

Reiter's Syndrome ◽

Reiter’S Syndrome

Diarrheic episodes caused by the foodborne pathogens Campylobacter, Salmonella, Shigella or Yersinia may lead to a sterile arthritis such as reactive arthritis, Reiter's syndrome or ankylosing spondylitis. Reiter's syndrome and reactive arthritis have been shown to be sequelae in a few well-studied bacterial food poisoning outbreaks. Reactive arthritis, Reiter's syndrome and ankylosing spondylitis show strong familial association related to the gene for HLA-B27 (HLA = human leucocyte antigen) antigen. Why HLA-B27-positive individuals are more susceptible to arthritis is not known, but molecular mimicry between the HLA-B27 antigen and antigens of triggering bacteria has been demonstrated and this mimicry has been proposed as a mechanism involved in etiology of the arthritides. Antigens from bacteria that triggered the arthritis are present in arthritic joints but bacterial cells are not found. Antibodies and T-cells specific for the triggering bacteria have been demonstrated in arthritic patients. T-cells present in synovial joints respond specifically to the particular arthritic triggering pathogen. The cells that respond to bacterial antigens belong to the T-cell subset TH1 that secrete a limited number of cytokines but it is not known if cytokines are involved in arthritis. A few studies have demonstrated that T-cells from the joints of arthritic patients respond to both bacterial and human heat shock proteins indicating that autoimmunity may be involved in causation of arthritis. While only about 2% of a population exposed to a triggering infection will acquire arthritis, these individuals undergo pain and suffering as well as economic hardships as a result of their disease.

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Overview of Plant-Derived Vaccine Antigens: Dengue Virus

Journal of Pharmacy & Pharmaceutical Sciences ◽

10.18433/j3401k ◽

2011 ◽

Vol 14 (3) ◽

pp. 400 ◽

Cited By ~ 7

Author(s):

Ravindra B Malabadi ◽

Advaita Ganguly ◽

Jaime A Teixeira da Silva ◽

Archana Parashar ◽

Mavanur R Suresh ◽

...

Keyword(s):

Infectious Diseases ◽

Dengue Virus ◽

Neutralizing Antibodies ◽

Vaccine Development ◽

Low Cost ◽

Current Status ◽

Human Immune Deficiency Virus ◽

Dengue Vaccine ◽

New Methods ◽

Vaccine Antigens

ABSTRACT - This review highlights the advantages and current status of plant-derived vaccine development with special reference to the dengue virus. There are numerous problems involved in dengue vaccine development, and there is no vaccine against all four dengue serotypes. Dengue vaccine development using traditional approaches has not been satisfactory in terms of inducing neutralizing antibodies. Recently, these issues were addressed by showing a very good response to inducing neutralizing antibodies by plant-derived dengue vaccine antigens. This indicates the feasibility of using plant-derived vaccine antigens as a low-cost method to combat dengue and other infectious diseases. The application of new methods and strategies such as dendritic cell targeting in cancer therapy, severe acute respiratory syndrome, tuberculosis, human immune deficiency virus, and malaria might play an important role. These new methods are more efficient than traditional protocols. It is expected that in the near future, plant-derived vaccine antigens or antibodies will play an important role in the control of human infectious diseases. This article is open to POST-PUBLICATION REVIEW. Registered readers (see “For Readers”) may comment by clicking on ABSTRACT on the issue’s contents page.

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High-Throughput Identification of New Protective Antigens from a Yersinia pestis Live Vaccine by Enzyme-Linked Immunospot Assay

Infection and Immunity ◽

10.1128/iai.00242-09 ◽

2009 ◽

Vol 77 (10) ◽

pp. 4356-4361 ◽

Cited By ~ 22

Author(s):

Bei Li ◽

Lei Zhou ◽

JingYu Guo ◽

Xiaoyi Wang ◽

Bin Ni ◽

...

Keyword(s):

T Cell ◽

Yersinia Pestis ◽

Cellular Immunity ◽

Vaccine Development ◽

Lethal Dose ◽

Live Vaccine ◽

Intracellular Bacteria ◽

Partial Protection ◽

Cell Responses ◽

Protective Antigens

ABSTRACT Yersinia pestis, the plague pathogen, is a facultative intracellular bacterium. Cellular immunity plays important roles in defense against infections. The identification of T-cell targets is critical for the development of effective vaccines against intracellular bacteria; however, the function of cellular immunity in protection from plague was not clearly understood. In this study, 261 genes from Y. pestis were selected on the basis of bioinformatics analysis and previous research results for expression in Escherichia coli BL21(DE3). After purification, 101 proteins were qualified for examination of their abilities to induce the production of gamma interferon in mice immunized with live vaccine EV76 by enzyme-linked immunospot assay. Thirty-four proteins were found to stimulate strong T-cell responses. The protective efficiencies for 24 of them were preliminarily evaluated using a mouse plague model. In addition to LcrV, nine proteins (YPO0606, YPO1914, YPO0612, YPO3119, YPO3047, YPO1377, YPCD1.05c, YPO0420, and YPO3720) may provide partial protection against challenge with a low dose (20 times the 50% lethal dose [20× LD50]) of Y. pestis, but only YPO0606 could partially protect mice from infection with Y. pestis at a higher challenge dosage (200× LD50). These proteins would be the potential components for Y. pestis vaccine development.

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Prokaryotic Single-Cell RNA Sequencing by In Situ Combinatorial Indexing

10.1101/866244 ◽

2019 ◽

Cited By ~ 3

Author(s):

Sydney B. Blattman ◽

Wenyan Jiang ◽

Panos Oikonomou ◽

Saeed Tavazoie

Keyword(s):

Single Cell ◽

Rna Sequencing ◽

Low Cost ◽

Bacterial Cells ◽

Single Experiment ◽

Bacterial Populations ◽

Single Cell Rna Sequencing ◽

Gene Expression Heterogeneity ◽

Mrna Polyadenylation

AbstractDespite longstanding appreciation of gene expression heterogeneity in isogenic bacterial populations, affordable and scalable technologies for studying single bacterial cells have been limited. While single-cell RNA sequencing (scRNA-seq) has revolutionized studies of transcriptional heterogeneity in diverse eukaryotic systems, application of scRNA-seq to prokaryotes has been hindered by their extremely low mRNA abundance, lack of mRNA polyadenylation, and thick cell walls. Here, we present Prokaryotic Expression-profiling by Tagging RNA In Situ and sequencing (PETRI-seq), a low-cost, high-throughput, prokaryotic scRNA-seq pipeline that overcomes these technical obstacles. PETRI-seq uses in situ combinatorial indexing to barcode transcripts from tens of thousands of cells in a single experiment. PETRI-seq captures single cell transcriptomes of Gram-negative and Gram-positive bacteria with high purity and low bias, with median capture rates >200 mRNAs/cell for exponentially growing E. coli. These characteristics enable robust discrimination of cell-states corresponding to different phases of growth. When applied to wild-type S. aureus, PETRI-seq revealed a rare sub-population of cells undergoing prophage induction. We anticipate broad utility of PETRI-seq in defining single-cell states and their dynamics in complex microbial communities.

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Characterization of enteric disease in children using a low-cost specimen preservation method

Journal of Clinical Microbiology ◽

10.1128/jcm.01703-21 ◽

2021 ◽

Author(s):

Amanda K. Debes ◽

Shaoming Xiao ◽

Jie Liu ◽

Allison Shaffer ◽

Paul Scalzo ◽

...

Keyword(s):

Filter Paper ◽

Disease Burden ◽

Stool Specimen ◽

Diarrheal Disease ◽

Vaccine Development ◽

Low Cost ◽

Preservation Method ◽

Stool Specimens ◽

Taqman Array

BACKGROUND: Diarrhea is a leading cause of death in children under five. Molecular methods exist for the rapid detection of enteric pathogens; however, the logistical costs of storing stool specimens limit applicability. We sought to demonstrate that dried filter paper specimen preservation can identify diarrheal diseases causing significant morbidity among children in resource constrained countries. METHODS: A sub-study was nested into cholera surveillance in Cameroon. Enrollment criteria included: enrollment between 8/1/16 - 10/1/18; age < 18 years; a stool specimen; ≥ three loose stools within 24hours with the presence of dehydration and/or blood. 7227 persons were enrolled, for which 2746 met enrollment criteria and 337 were included in this analysis using the enteric TaqMan Array Card. Bacterial pathogens were compared to severity of diarrhea, age and sex, among other variables. RESULTS: 107 were ETEC positive of which: 40.2% (N=43) LT-STh, 19.6% (N=21) LT-STp; and 49.5% (53) LT-only. Major CFs were present in 43.9% of ETEC-positives. 96 were positive for Shigella, of which 14 (14.6%) reported dysentery. Model-derived quantitative cutoffs identified 116 (34.4%) with one highly diarrhea-associated pathogen and 16 (4.7%) with ≥ two. Shigellae and rotavirus were most strongly associated with diarrhea in children with mixed infections. CONCLUSION: Dried filter paper preserved specimens eliminate the need for frozen stool specimens and will facilitate enteric surveillance and contribute to the understanding of disease burden, which is needed to guide vaccine development and introduction. This study confirms Rotavirus, Shigella and ETEC as major contributors to pediatric diarrheal disease in two regions of Cameroon.

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LOW ENERGY BACTERIA PRESERVATION OF EXTREMELY HALOPHILIC ARCHAEA HALOFERAX LUCENTENSE AND HALOFERAX CHUDINOVII IMMOBILIZED USING NATURAL ZEOLITE

Jurnal Riset Teknologi Pencegahan Pencemaran Industri ◽

10.21771/jrtppi.2019.v10.no2.p16-28 ◽

2019 ◽

Vol 10 (2) ◽

pp. 16-28

Author(s):

Rizal Awaludin Malik ◽

Nilawati Nilawati ◽

Novarina Irnaning Handayani ◽

Rame Rame ◽

Silvy Djayanti ◽

...

Keyword(s):

Surface Area ◽

Pore Volume ◽

Low Cost ◽

Halophilic Archaea ◽

High Energy ◽

Slight Reduction ◽

Bacterial Cells ◽

Preservation Method ◽

Chemical Content ◽

Physical And Chemical

The methods of microbial cells preservation were already known by liquid drying, freeze-drying, and freezing. Those methods could preserve bacteria cells in a long period of time but its survivability was relatively low and used relatively high energy during preservation. Immobilization was known as entrapping, attaching or encapsulating bacterial cells in a suitable matrix. This research was conducted to know the suitability of zeolite as immobilization carrier and also as preservation matrix of two halophilic archaea Haloferax chudinovii and Haloferax lucentense. Variable of this research was the type of the carrier which was raw zeolite, 110oC and 300oC heat-activated zeolite carrier, parameters measured in this study was physical and chemical of zeolite such as chemical content, Si/Al ratio, surface area and pore volume, and biochemical assay, bacterial cells numbers after immobilization and bacterial cells after preservation as bacterial response to the immobilization and preservation. Heat activation was significantly affecting the chemical composition, carrier surface area, and pore volume. Highest surface area, pore volume, and Si/Al ratio were obtained in 110oC pretreated zeolite followed by 300oC pretreated zeolite. The bacterial cells obtained after immobilization process was 1,8x107 cfu/g, 3,0 x 107 cfu/g, and 2,1x107 for raw zeolite, 110oC pretreated zeolite and 300oC zeolite respectively. After 4 months preservation, the slight reduction of the bacterial cells was observed. Immobilization halophilic archaeae using zeolite as carrier was proven as low cost and effective preservation method due to relatively simple process and unspecific preservation temperature requirements.

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Bacterial Cell Inactivation Using a Single-Frequency Batch-Type Ultrasound Device

Indonesian Journal of Science and Technology ◽

10.17509/ijost.v6i1.31516 ◽

2021 ◽

Vol 6 (1) ◽

pp. 65-80

Author(s):

Poetro Sambegoro ◽

Maya Fitriyanti ◽

Bentang Arief Budiman ◽

Kamarisima Kamarisima ◽

Sekar Wangi Arraudah Baliwangi ◽

...

Keyword(s):

Bacterial Cell ◽

Treatment Time ◽

Low Cost ◽

Single Frequency ◽

Bacterial Cells ◽

Cell Inactivation ◽

E Coli ◽

Commercial System ◽

Ultrasound Device ◽

Frequency Ultrasound

Ultrasound technology employs cavitation to generate high-pressure soundwaves to disrupt bacterial cells. This study reveals the effectiveness of a single frequency ultrasound device for bacterial cell inactivation. A low-cost ultrasound device having a single frequency, i.e. 22 kHz for lab-scale application, was developed first, and the prototype was mechanically designed and analyzed using the finite-element method to assure the targeted natural frequency could be achieved. The prototype was then tested inactivating bacterial cells, Escherichia coli (E. coli) and Bacillus subtilis (B. subtilis), in a simple medium and a food system, and the results were then compared to a commercial system. A treatment time of up to 15 minutes was able to reduce E. coli and B. subtilis cells by 3.3 log and 2.8 log, respectively, and these results were similar to those of the commercial system. The effectiveness of bacterial cell inactivation using the developed single-frequency ultrasound device is then discussed. The findings are useful for designing low-cost ultrasound devices for application in the food industry.

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Low-cost management of mushroom poisoning in a limited-resource area: a 12-year retrospective study

Tropical Doctor ◽

10.1177/0049475519897704 ◽

2020 ◽

Vol 50 (2) ◽

pp. 135-138

Author(s):

Ratsameekhae Jongthun ◽

Pasin Hemachudha ◽

Supaporn Wacharapluesadee ◽

Thiravat Hemachudha

Keyword(s):

Medical Records ◽

Cost Management ◽

Low Cost ◽

Food Poisoning ◽

Alcoholic Cirrhosis ◽

Limited Resource ◽

Late Presentation ◽

Mushroom Poisoning ◽

Accidental Ingestion ◽

Poisonous Mushrooms

Amatoxin poisoning is the main cause of death from accidental ingestion of poisonous mushrooms and a mortality rate of 27.3% has been reported in Thailand. Symptoms of mushroom ingestion are often confused with food poisoning; thus, gastroenteritis is not recognised as the first phase of poisoning. Our study assessed the efficacy of N-acetylcysteine (NAC) as a treatment for amatoxin poisoning. We retrospectively analysed 74 medical records over 12 years. The majority (70/74) were treated successfully with NAC; death in the remaining 4 (5.4%) patients was attributed to late presentation in three and advanced alcoholic cirrhosis in one.

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