Magdy, H., M. H. Rady, M. S. Salama, H. A. E. Sayed, D. Hamza, M. Azzam, and E. E. Essa, "Isolation of Multidrug-Resistant from Wild Houseflies with a New Perspective for the Treatment.", Vector borne and zoonotic diseases (Larchmont, N.Y.), vol. 23, issue 2, pp. 63-74, 2023. Abstract

High frequency of infection and the unknown mode of transmission prompted us to investigate -wild housefly relationship. causes chronic gastritis, peptic ulcers, and stomach cancer. persists in the gut of the experimentally infected houseflies. The existence of strains isolated from wild houseflies, on the other hand, has never been documented. In this study, 902 wild houseflies from different sites were identified as , then 60 flies were screened by traditional microbiological techniques and -specific gene. The antibiotic resistance (ART) was investigated phenotypically. Wild housefly gut bacterial isolates were further evaluated genotypically to have gene mutation related to clarithromycin resistance. To find efficient therapeutic alternatives, the potency of three plant extracts (garlic, ginger, and lemon) and the wasp, venom was evaluated against . The cytotoxic effect of the crude wasp venom, the most potent extract, against Vero and Colon cancer (Caco2) cell lines was investigated using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. All isolates from houseflies were positive. The isolated bacteria have variable resistance to frequently used antibiotics in all isolates. Minimum inhibitory concentration values of 15.625 mg/mL for both ginger and lemon extracts, 7.8125 mg/mL for garlic extract, and 0.0313 mg/mL for wasp venom were recorded. Wasp venom has the most potent antibacterial activity compared with the four antibiotics that are currently used in therapies against . We conclude that wild houseflies can play a role in disseminating The housefly gut may be a suitable environment for the horizontal transfer of ART genes among its associated microbiome and Wasp venom proved its potential activity as a new and effective anti- drug for both therapeutic and preventative usage.

Mario, E., D. Hamza, and K. Abdel-Moein, "Hypervirulent Klebsiella pneumoniae among diarrheic farm animals: A serious public health concern.", Comparative immunology, microbiology and infectious diseases, vol. 102, pp. 102077, 2023. Abstract

Hypervirulent Klebsiella pneumoniae (hvKp) is an emerging pathogen and it has more virulence factors than classical Klebsiella pneumoniae strains. Carbapenem-resistant hvKp (CR-hvKp) is a dangerous bacteria that has both high virulence and antibiotic resistance and poses a global public health problem worldwide. The current study was carried out to investigate the occurrence of hvKp as well as carbapenem-resistant Klebsiella pneumoniae (CRKP) and CR-hvKp among diarrheic farm animals. For this purpose, rectal swabs from 165 farm animals (45 cattle, 66 sheep, and 54 goats) were collected. Samples were processed for the isolation and identification of Klebsiella pneumoniae. Moreover, hvKp was detected using molecular techniques by amplification of biomarker virulence genes (rmpA, rmpA2, iucA, iroB, and peg-344), followed by a string test. On the other hand, all K. pneumoniae isolates were examined for carbapenem resistance by both phenotypic and molecular methods. The phylogenetic analysis of peg-344 sequences was carried out. The overall prevalence rates of K. pneumoniae, hvKp, CRKP, and CR-hvKp were 24.2%, 7.9%, 16.4%, and 6.1% respectively. HvKp and CR-hvKp were detected among all examined farm animal species. On a Molecular basis, all biomarker virulence genes were identified except iroB, but rmpA is the most prevalent one. The phylogenetic analysis of peg-344 sequences obtained from the study points out their genetic relatedness to those circulated among humans. In conclusion, the emergence of hvKp and CR-hvKp among diarrheic farm animals confers a great public health implication and thus, the possible animal reservoirs for such hypervirulent-antimicrobial resistant strains cannot be ruled out.

Bhattacharya, K., I. M. Shamkh, M. S. Khan, M. M. Lotfy, J. B. Nzeyimana, R. F. Abutayeh, N. M. Hamdy, D. Hamza, N. R. Chanu, P. Khanal, et al., "Multi-Epitope Vaccine Design against Monkeypox Virus via Reverse Vaccinology Method Exploiting Immunoinformatic and Bioinformatic Approaches.", Vaccines, vol. 10, issue 12, 2022. Abstract

(1) Background: The monkeypox virus is a zoonotic orthopox DNA virus that is closely linked to the virus. In light of the growing concern about this virus, the current research set out to use bioinformatics and immunoinformatics to develop a potential vaccine against the virus. (2) Methods: A multiepitope vaccine was constructed from the B-cell and T-cell epitopes of the MPXVgp181 strain using adjuvant and different linkers. The constructed vaccine was predicted for antigenicity, allergenicity, toxicity, and population coverage. In silico immune simulation studies were also carried out. Expression analysis and cloning of the constructed vaccine was carried out in the pET-28a(+) vector using snapgene. (3) Results: The constructed vaccine was predicted to be antigenic, non-allergenic, and non-toxic. It was predicted to have excellent global population coverage and produced satisfactory immune response. The in silico expression and cloning studies were successful in , which makes the vaccine construct suitable for mass production in the pharmaceutical industry. (4) Conclusion: The constructed vaccine is based on the B-cell and T-cell epitopes obtained from the MPXVgp181 strain. This research can be useful in developing a vaccine to combat the monkeypox virus globally after performing in-depth in vitro and in vivo studies.

Mohammed, R., S. M. Nader, D. A. Hamza, and M. A. Sabry, "Horse: a potential source of Cryptococcus neoformans and Cryptococcus gattii in Egypt.", BMC veterinary research, vol. 18, issue 1, pp. 17, 2022. Abstract

BACKGROUND: Cryptococcosis is an opportunistic mycozoonosis of global significance in a wide variety of host species. In equines, cryptococcosis is uncommon, and sporadic cases have been reported with rhinitis, sinusitis, pneumonia, and meningitis. Cryptococcus spp. represents a potential risk for immunosuppressed and healthy persons. In Egypt, epidemiological data on cryptococcal infection in horses are limited. The current study was carried out to investigate the occurrence of Cryptococcus spp. in horses and its possible role in the epidemiology of such disease in Egypt. A total of 223 samples was collected from different localities in Egypt included 183 nasal swabs from horses, 28 nasal swabs from humans, and 12 soil samples. Bacteriological examination and the identification of Cryptococcus spp. were performed. Molecular serotyping of Cryptococcus spp. was determined by multiplex PCR using CNa-70S/A-CNb-49S/A. The virulence genes (LAC1, CAP59, and PLB1) of the identified isolates were detected by PCR. Moreover, sequencing and phylogenetic analysis of the C. gattii gene from horses, humans, and soil isolates found nearby were performed.

RESULT: The overall occurrence of Cryptococcus spp. in horses were 9.3, 25, and 10.7% in horses, the soil, and humans, respectively. Molecular serotyping of the Cryptococcus spp. isolates recovered from the nasal passages of horses proved that C. gattii (B), C. neoformans, and two hybrids between C. neoformans (A) and C. gattii (B) were identified. Meanwhile, in case of soil samples, the isolates were identified as C. gattii (B). The human isolates were serotyped as C. gattii in two isolates and C. neoformans in only one isolate. Molecular detection of some virulence genes (LAC1), (CAP59), and (PLB1) were identified in both C. gattii and C. neoformans isolates. The C. gattii gene amplicons of the isolates from horses, humans, and the soil were closely related.

CONCLUSION: This study provides the first insights into the Egyptian horse ecology of Cryptococcus species and highlights the role of horses as asymptomatic carriers in disseminating the potentially pathogenic Cryptococcus spp. It also presents the possible risk of cryptococcosis infection in humans.

Elkazzaz, M., A. Ahmed, Y. E. - E. Abo-Amer, T. Hydara, A. Haikal, D. A. E. N. Razek, W. A. Eltayb, X. Wang, T. M. Karpiński, D. Hamza, et al., "In Silico Discovery of GPCRs and GnRHRs as Novel Binding Receptors of SARS-CoV-2 Spike Protein Could Explain Neuroendocrine Disorders in COVID-19.", Vaccines, vol. 10, issue 9, 2022. Abstract

Despite the intense research work since the beginning of the pandemic, the pathogenesis of COVID-19 is not yet clearly understood. The previous mechanism of COVID-19, based on ACE2 tropism and explained through a single receptor, is insufficient to explain the pathogenesis due to the absence of angiotensin-converting enzyme 2 (ACE2) receptors in most of the affected organs. In the current study, we used the PatchDock server to run a molecular docking study of both the gonadotropin-releasing hormone receptor (GnRHR) and G-protein-coupled-receptor (GPCR) with the SARS-CoV-2 spike protein. Molecular Dynamics (MD) simulations were run to analyze the stability of the complexes using the GROMACS package. The docking results showed a high affinity between the spike protein with the GnRHR (-1424.9 kcal/mol) and GPCR (-1451.8 kcal/mol). The results of the MD simulations revealed the significant stability of the spike protein with the GnRHR and GPCR up to 100 ns. The SARS-CoV-2 spike protein had strong binding interactions with the GPCRs and GnRHRs, which are highly expressed in the brain, endocrine organs, and olfactory neurons. This study paves the way towards understanding the complex mechanism of neuroendocrine involvement and peripheral organ involvement, may explain the changing symptoms in patients due to new variants, and may lead to the discovery of new drug targets for COVID-19. In vitro studies involving genetic engineering or gene knockdown of the GPCRs and GnRHRs are needed to further investigate the role of these receptors in COVID-19 pathogenesis.

Elhariri, M., R. Elhelw, S. Selim, M. Ibrahim, D. Hamza, and E. Hamza, "Virulence and Antibiotic Resistance Patterns of Extended-Spectrum Beta-Lactamase-Producing serovar Heidelberg Isolated from Broiler Chickens and Poultry Workers: A Potential Hazard.", Foodborne pathogens and disease, vol. 17, issue 6, pp. 373-381, 2020. Abstract

The current study investigated the emergence of multidrug-resistance (MDR), extended-spectrum beta-lactamase (ESBL)-producing serovar Heidelberg in broiler chickens and workers in poultry farms. A total of 33 . Heidelberg isolates were recovered; 24 from the broiler cloacal swabs and 9 from the farm workers. All the . Heidelberg isolates were tested for susceptibility to 11 antimicrobial agents and for the presence of resistance and virulence genes. MDR strains were found in 95.8% (23/24) and 88.8% (8/9) of the broiler and human isolates, respectively. Among the MDR strains, 66.6% of the broiler isolates and 55.5% of the human isolates were ESBL producing. The majority of broiler isolates showed resistance to ampicillin (100%) and ceftriaxone (91.6%), followed by ceftazidime and imipenem, (87.5%) and (75%). The resistance rate of the human isolates to those antibiotics were lower than the broiler isolates; ampicillin (88.8%), ceftriaxone (66.6%), ceftazidime (77.7%), and imipenem (66.6%). The resistance determinant genes found among the isolated strains was , , , , , , and . The most detected ESBL genes for broiler and human isolates were (63.7%) and (56.6%), followed by (48.5%), (39.4%), and (27.3%); whereas and were not detected. The finding of chromosomal and plasmid virulence genes revealed that the A (100%), , C, and (72.8%), C (66.7%), (63.6%), B (54.6%), and A and A (3.0%), while A and R were absent. An elevated rate of MDR Heidelberg in chickens is of potential great health risk. This signifies the role of the food of animal origin as a reservoir of MDR that can affect the human health.

Hamza, D., S. Dorgham, E. Ismael, S. I. A. El-Moez, M. Elhariri, R. Elhelw, and E. Hamza, "Emergence of β-lactamase- and carbapenemase- producing Enterobacteriaceae at integrated fish farms.", Antimicrobial resistance and infection control, vol. 9, issue 1, pp. 67, 2020. Abstract

BACKGROUND: Epidemiological studies suggested that determinants for antibiotic resistance have originated in aquaculture. Recently, the integrated agriculture-aquaculture system has been implemented, where fish are raised in ponds that receive agriculture drainage water. The present study aims to investigate the occurrence of β-lactamase and carbapenemase-producing Enterobacteriaceae in the integrated agriculture-aquaculture and the consequent public health implication.

METHODS: Samples were collected from fish, fishpond water inlets, tap water, outlet water, and workers at sites of integrated agriculture-aquacultures. Samples were also taken from inhabitants of the aquaculture surrounding areas. All samples were cultured on MacConkey agar, the Enterobacteriaceae isolates were tested for susceptibility to cephalosporins and carbapenems, and screened for bla, bla, bla, bla, bla, bla, bla, and bla. Strains having similar resistance phenotype and genotype were examined for the presence of Incompatible (Inc) plasmids.

RESULTS: A major proportion of the Enterobacteriaceae isolates were resistant to cephalosporins and carbapenems. Among the 66 isolates from fish, 34 were resistant to both cephalosporin and carbapenem groups, 26 to carbapenems alone, and 4 to cephalosporins alone. Of the 15 isolates from fishpond water inlets, 8 showed resistance to both groups, 1 to carbapenems alone, and 5 to cephalosporins alone. Out of the 33 isolates from tap water, 17 were resistant to both groups, and 16 to cephalosporins alone. Similarly, of the 16 outlet water isolates, 10 were resistant to both groups, and 6 to cephalosporins alone. Furthermore, of the 30 examined workers, 15 carried Enterobacteriaceae resistant strains, 10 to both groups, and 5 to cephalosporins alone. Similar strains were isolated from the inhabitants of the aquaculture surrounding areas. Irrespective of source of samples, strains resistant to all examined antibiotics, carried predominantly the carbapenemase gene bla either alone or with the β-lactamase genes (bla, bla, bla, and bla). The isolates from fish, water, and workers harboured a wide-range of multi-drug-resistance Inc. plasmids, which were similar among all isolates.

CONCLUSION: The present findings suggest transmission of the resistance genes among Enterobacteriaceae strains from different sources. This reiterates the need for control strategies that focus on humans, animals, water, and sewage systems to solve the antibiotic resistance problem.

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