Development, Optimization and Evaluation of a Sensitive Enzyme-Linked Immunosorbent Assay (ELISA) Prototype for Detection of Chicken-Based IgY Polyclonal Antibodies against Toxins of D. polylepis Venom.

Life-threatening medical issues can result from snakebite, and hence this is a public health concern. In many tropical and subtropical nations such as Kenya, where a wide variety of poisonous snakes are prevalent, diagnosis of snakebite in health facilities is imperative. Different antivenoms are needed to treat the venom of different snake species. Nonetheless, it might be difficult for medical professionals to identify the exact snake species that envenomated a patient due to the similarities of several snake envenomations' clinical symptoms. Therefore, the necessity for an assay or technique for identifying venomous species is critical. The current study sought to develop a sensitive ELISA prototype for the detection of D. polylepis venom in Kenya using generated chicken-based IgY polyclonal antibodies. Serum samples containing specific chicken-based IgY antibodies previously raised against D. polylepis venom toxins were used in the assay development. ELISA parameters were optimized, and the developed assay was assessed for applicability. The limit of detection (LoD) of the ELISA for neurotoxic venoms was determined to be 0.01 µg/mL. Successful discrimination between neurotoxic and cytotoxic venoms was achieved by the ensuing inhibition ELISA assay. The developed assay showed the capability of identifying venoms in blood samples (from spiked and venom-challenged blood samples) of BALB/c mice, providing compelling evidence of the strategy's usefulness. This assay could help physicians diagnose and manage victims of snakebites through the evaluation of clinical samples.

Generation of chicken-based IgY polyclonal antibodies against Dendroaspis polylepis and preclinical evaluation of envenomation-neutralizing efficacy vis-à-vis selected commercial antivenoms.

The Black mamba, D. polylepis, is one of the many venomous snakes found in Kenya, and known to account for some snakebite incidents. The Kenyan Ministry of Health data reveals annual 15,000 snakebites occurrences. Also, 1 in 15 people in Kenya gets bitten by a snake, and tragically, 1 in 147 of these individuals die of snakebite yearly. Traditionally, antivenoms for treatment are produced from horse or sheep but have complicated and expensive production issues. Alternative production approaches, such as using IgY antibodies derived from chicken egg yolks, may overcome disadvantages with traditional antivenom manufacturing techniques. In this current study, D. polylepis specific IgY polyclonal antibodies were purified from the egg yolks of chickens immunized with D. polylepis venom. These antibodies were subsequently assessed for their in-vivo neutralizing capacity vis-à-vis commercial antivenoms, PANAF-Premium and VINS. The IgY antibodies were purified by ammonium sulfate precipitation and affinity-chromatography, with quality and specificity determined by SDS-PAGE and ELISA. The LD50 of D. polylepis was found to be 0.54 mg/kg in chicks, and 0.34 mg/kg in mice, respectively. Pool of extracted IgY yielded 2.8 mg/mL concentration. Purified IgY under non-reducing and reducing conditions on SDS-PAGE exhibited a single-protein band of about 183 kDa and two bands (67 kDa and 25 kDa), respectively. The minimum-edematogenic dose was 0.05 µg. Anti-D. polylepis IgY antibodies and two antivenoms demonstrated the capacity to neutralize the toxic activities of D. polylepis venom. This study confirms a successful IgY generation against Black mamba venom for the first time, and observed toxic effects of the venom as well as neutralizing capacity of antivenoms.

Antibiotic resistance pattern of methicillin-resistant Staphylococcus aureus and Escherichia coli from mobile phones of healthcare workers in public hospitals in Ghana.

Introduction: mobile phone plays an essential role in the lives of healthcare professionals in hospitals as far as communication is concerned. However, it can also serve as a source of nosocomial infections. This study aimed at determining the prevalence and antibiotic susceptibility of Methicillin-resistant Staphylococcus aureus (MRSA) and Escherichia coli isolated from mobile phones used by healthcare staff working in three public hospitals in Ghana. Methods: in total, 220 swab samples were collected from 110 mobile phones of healthcare workers at a referral and two public tertiary hospitals in Ghana. Direct spreading of swab samples on agar plates was done. MacConkey agar and Baird Parker agar were used to isolate E. coli and S. aureus, respectively. Clinical Laboratory Standard Institute´s guidelines were followed for susceptibility testing, and S. aureus strains resistant to cefoxitin were considered to be MRSA. All E. coli and MRSA isolates were tested for their susceptibility to antibiotics using European Committee on Antimicrobial Susceptibility Testing (EUCAST) 2018 guidelines with its breakpoints. Obtained qualitative data were analyzed by using Microsoft Excel. Results: of 110 mobile phones, 78 (70.9%) and 4 (3.6%) were colonized with S. aureus and E. coli, respectively. From the 78 S. aureus isolates, 22 (28%) isolates were MRSA. Fifty percent (50%) (11/22) of the MRSA isolates were multi-drug resistant, of which one isolate was resistant to all antibiotics tested. E. coli isolates had 100 resistances to both ceftriaxone and ceftazidime. Conclusion: mobile phones used by healthcare workers in hospitals frequently harbor E. coli, S. aureus, MRSA and may be sources of hospital-associated infections.

TOOTHBRUSH AND TOWEL HANDLING AND THEIR MICROBIAL QUALITY: THE CASE OF STUDENTS OF UNIVERSITY FOR DEVELOPMENT STUDIES, NYANKPALA CAMPUS, GHANA.

BACKGROUND: Good toothbrush and towel handling are important considerations in personal hygiene. Thus, this study sought to assess how students of the University for Development Studies handle their toothbrushes and towels and the consequence of that with regards to the microbial quality of these personnel hygiene materials. MATERIALS AND METHODS: A total of 100 swap samples were collected (50 toothbrushes and 50 towels) for microbial analysis. Questionnaires were administered to students from whom samples were collected to ascertain information on how they handle toothbrushes and towels. MacConkey agar and Mannitol Salt agar were used to isolate E. coli and S. aureus respectively, and cefoxitin used to identify the methicillin-resistant S. aureus strains. RESULTS: E. coli was present in all sampled towels, while 98% of the sampled toothbrushes contained E. coli. It was found that 2% of the respondents kept their toothbrushes in bathhouses, 44% kept them unenclosed in rooms and 54% kept them enclosed in rooms (54%). Also, 48% of the respondents washed their towels once a week, 24% washed once every two weeks, 20% once every month and 8% once a trimester. Moreover, 52% dried their towels in rooms while 48% dried them outside rooms. The occurrence of S. aureus was 96% and 94% respectively for the towels and toothbrushes. Of the S. aureus isolated, 33.3% of sampled towels and 12.8% of the toothbrushes contained methicillin-resistant S. aureus. CONCLUSION: This study found that, students are at risk of contracting infectious disease if their personal hygiene behaviours do not changed.

Prevalence and pattern of antibiotic resistance of Staphylococcus aureus isolated from door handles and other points of contact in public hospitals in Ghana.

BACKGROUND: Studies have implicated Staphylococcus aureus as the leading cause of septicemia in the Tamale metropolis of Ghana. The aim of this study was to determine the prevalence and antibiotic susceptibility of S. aureus and Methicillin Resistant S. aureus (MRSA) in the environments of three hospitals in Ghana. METHODS: A total of 120 swab samples were taken from door handles, stair railings and other points of contact at Tamale Teaching Hospital, Tamale Central Hospital and Tamale West Hospital. The swab samples were directly plated on Mannitol Salt and Baird Parker agar plates and incubated at 37 °C (± 2) for 18-24 h. An antibiotic susceptibility test was performed using the Clinical Laboratory Standard Institute's guidelines. Isolates resistant to both cefoxitin and oxacillin were considered to be MRSA. RESULTS: A total of 47 (39%) positive S. aureus samples were isolated from all three hospitals, of which, eight (17%) were putative MRSA isolates. One MRSA isolate was resistant to all the antibiotics used (cefoxitin, oxacillin, ciprofloxacin, erythromycin, tetracycline, ampicillin, streptomycin and sulfamethoxazole-trimethoprim). Five of the MRSA isolates were multi-drug resistant, whilst the other three were resistant to only two antibiotics. All the multi-drug resistant MRSA isolates were resistant to at least four antibiotics. The percentage of isolates resistant to oxacillin, ampicillin, ciprofloxacin, tetracycline, streptomycin, erythromycin, and sulfamethoxazole/trimethoprim were 17, 13, 9, 28, 89, 13 and 11% respectively. CONCLUSION: The high multi-drug resistance of MRSA in hospital environments in Ghana reinforces the need for the effective and routine cleaning of door handles in hospitals. Further investigation is required to understand whether S. aureus from door handles could be the possible causes of nosocomial diseases in the hospitals.