In vivo experimentally study the effect of Nigella Sativa silver nanoparticles for treatment of salmonella species causing diarrhea in ruminants.

Salmonellais one of the main etiological agents of infectious diarrhea in large and small ruminants but emergence of multidrug-resistant (MDR) strains faster rate than previously, leads to develop of MDR strains among animals needs different alternative therapeutic strategies. Our study was aimed to evaluate the effects of Nigella sativa silver nanoparticles (NS AgNPs) on specific pathogen-free (SPF) Wister rats. Nigella sativa silver nanoparticles were prepared and confirmed their formation by optical observations, UV-Vis spectroscopy, transmission electron microscopy (TEM), and scanning electron microscopy (SEM). Rats in group G2 were infected experimentally with Salmonella spp and treated with ciprofloxacin orally for duration of 6 days at a dose rat 10 mg/kg. On the other hand, rats in group G1 were infected with salmonella and treated for 20 days with NS AgNPs in oral dose of (10 mg/kg rats), and the results were compared to control groups G3 which received bacterial infection without treatment and G4 control negative. The results of optical observation, UV-Vis spectroscopy, TEM, and SEM revealed typical characteristics of prepared NS AgNPs. Liver, kidney function biomarkers, hematologic analysis, and histological examination the tissues of liver, kidney, and stomach of rat's model improved that NS AgNPs has antimicrobial effect and has the ability to decrease the inflammatory reaction caused by Salmonella spp infection. The results of our study indicate that NS AgNPs are effective in controlling MDR Salmonella spp in vivo without causing any adverse effects. Moreover, our findings suggest that reducing the use of antimicrobials could be a key factor in the fight against antimicrobial resistance and can provide valuable insights into identifying the most appropriate treatment strategies to tackle this issue effectively in the future.

Advances in diagnosis and control of anthelmintic resistant gastrointestinal helminths infecting ruminants.

Infection with gastrointestinal helminths is widely spread among ruminant causing severe losses and adversely affects the livestock husbandry. Synthetic chemotherapeutics have been utilized throughout years, as a means of combating helminthiasis. Anthelmintic resistance (AR) has a serious concern on livestock industry which, mainly arises as outcome of misuse, improper dosing and frequent utilization of the synthetic drugs.Various gastrointestinal helminths have the capability to survive the therapeutic dose of anthelmintics and become resistant to the major anthelmintic classes. Early diagnosis might delay or reduce the risk of AR. Conventional phenotyping methods were commonly used for detection of anthelmintic resistant helminths, but appeared to lack of sensitivity, especially when the frequency of resistant allele is very low. Several molecular assays were carried out to detect the AR with greater accuracy. Sustainable effective preventive and control measures for gastrointestinal helminths infection remain the corner stone to overcome AR. Rational use of anthelmintics with keeping unexposed proportion of worm populations, could have the potentiality to maintain and prolong the efficacy of anthelmintics. Several alternative anthelmintic treatments might offer valuable solutions either alone or adjunct to synthetic drugs to dilute the spread of resistance alleles among the helminths population. This article reviews current status of various diagnostic methods and control measures for anthelmintic resistant gastrointestinal helminths infecting ruminants and tries to present a practical protocol to avoid or delay the development of AR.

Therapeutic effect of biosynthetic gold nanoparticles on multidrug-resistant Escherichia coli and Salmonella species isolated from ruminants.

BACKGROUND AND AIM: Multidrug-resistant (MDR) pathogenic microorganisms have become a global problem in ruminants as a result of the intensive use of antibiotics, causing the development of resistance among gut microbiota. The antibiotic-resistant microorganisms can be transferred from diseased animals to humans. This study aimed to determine the prevalence of MDR Escherichia coli and Salmonella spp. isolated from cattle, buffaloes, sheep, and goats suffering from respiratory signs, diarrhea, and mastitis and to screen the antibiotic sensitivity of selected isolated bacteria. It also detected antibiotic-resistance genes by polymerase chain reaction (PCR), produced green gold nanoparticles (AuNPs) using plant extracts (Artemisia herba-alba and Morus alba), and evaluated the antimicrobial activities of these biosynthesized nanoparticles on selected pathogens (E. coli and Salmonella spp.). MATERIALS AND METHODS: MDR E. coli and Salmonella spp. were investigated using fecal samples (n=408), nasal swabs (n=358), and milk samples (n=227) of cattle, buffaloes, sheep, and goats with or without clinical signs, including respiratory manifestations, pneumonia, diarrhea, and mastitis, from different governorates in Egypt. E. coli and Salmonella spp. were isolated and identified on selective media, which were confirmed by biochemical reactions and PCR. Antimicrobial susceptibility testing against 10 commonly used antibiotics was performed using the Kirby-Bauer disk diffusion method. Antibiotic resistance genes blaTEM, blaSHV, blaOXA , and bla CTX-M were detected by PCR. The antibacterial effect of the biosynthesized AuNPs was evaluated by MIC and well diffusion assay. The biosynthesized AuNPs were also characterized by ultraviolet-visible spectrophotometry and transmission electron microscopy (TEM). RESULTS: Among all fecal samples, the prevalence of E. coli was 18.4% (183/993) and that of Salmonella spp. was 16.7% (66/408), as determined by cultural and molecular tests. All isolates of E. coli and Salmonella spp. were 100% resistant to ampicillin (AM) and amoxicillin and highly resistant to cefoxitin and AM-sulbactam. The total rate of resistance genes in E. coli was 61.2% (112/183), while that in Salmonella was 63.6% (42/66) for pathogens isolated from ruminants with respiratory manifestations, pneumonia, diarrhea, and mastitis. Among the resistance genes, blaTEM had the highest prevalence rate in E. coli (25.9%, 21/81) while blaSHV had the lowest (9.8%, 8/81) in fecal swabs. AuNPs were successfully synthesized using aqueous leaf extract of A. herba-alba and M. alba as bioreducing agents. TEM analysis showed particle size of 10-42 nm for A. herba-alba and M. alba AuNPs. The biosynthesized AuNPs showed antibacterial activity against MDR E. coli and Salmonella spp. CONCLUSION: Rapid and accurate diagnostic methods are the cornerstone for effective treatment to reduce the risk of antimicrobial-resistant pathogenic microorganisms. This is particularly important for overcoming the increasing rate of MDR in ruminants with respiratory manifestations, pneumonia, diarrhea, and mastitis. This can be complemented by the development of AuNPs synthesized in an environmentally friendly manner AuNPs using natural plant extracts for the treatment of antibiotic-resistant microorganisms.

Development and characterization of ORF68 negative equine herpes virus type-1, Ab4p strain.

Equine herpesvirus-1 (EHV-1) is an important pathogen, which infects horses worldwide with high morbidity but low mortality rates. The respiratory disorders and abortions are the most common indicators. Ab4p (an abortigenic and paralytic virus) is one of the most important and virulent strains. The development and functional characterization of the open reading frame-68 (ORF68) negative EHV-1 Ab4p mutants and an assessment of their roles in the infection at the cellular level were the main targets of the current study. Escherichia coli DH10ß containing the Ab4p bacterial artificial chromosome (pAb4pBAC) and Red/ET expression vector were used to develop different ORF68 mutants. Multi-step growth kinetic experiments were conducted in order to evaluate the growth properties of the constructed mutant viruses. Growth of the Ab4pΔORF68 showed the lowest titer, compared to the Ab4pΔORF68R, Ab4pΔORF68R non-sense, and the parent Ab4p viruses without any significant difference (P > 0.05). The growth of the mutant viruses was almost similar across the cell types, but viruses growth was more efficient in FHK cells as judged by the number of the obtained virus particles. The plaque size of Ab4pΔORF68 was significantly (40%) smaller than those of Ab4p (P < 0.01), Ab4pΔORF68R, and Ab4pΔORF68R non-sense viruses which confirmed the importance of ORF68 protein in the cell-to-cell transmission of EHV-1. Subcellular localization of the green fluorescent protein (GFP) ORF68 gene fusion product showed late expression with intranuclear localization of the transfected cells while immunofluorescent antibody technique (IFAT) localized it at the nucleus and nuclear membranes of the infected cells. Hence, it could be concluded that ORF68 protein may not be essential for EHV-1 Ab4p growth but plays a crucial role in virus penetration and transmission at the cellular level. Therefore, the generated EHV-1 ORF68 negative mutant could be a prospective candidate for the development of a vaccine marker.

Morphological and molecular identification of the brown dog tick Rhipicephalus sanguineus and the camel tick Hyalomma dromedarii (Acari: Ixodidae) vectors of Rickettsioses in Egypt.

AIM: Rickettsioses have an epidemiological importance that includes pathogens, vectors, and hosts. The dog tick Rhipicephalus sanguineus and the camel tick Hyalomma dromedarii play important roles as vectors and reservoirs of Rickettsiae. The aim of this study was to determine the prevalence of Rickettsiae in ixodid ticks species infesting dogs and camels in Egypt, in addition to, the morphological and molecular identification of R. sanguineus and H. dromedarii. MATERIALS AND METHODS: A total of 601 and 104 of ticks' specimens were collected from dogs and camels, respectively, in Cairo, Giza and Sinai provinces. Hemolymph staining technique and OmpA and gltA genes amplification were performed to estimate the prevalence rate of Rickettsiae in ticks. For morphological identification of tick species, light microscope (LM) and scanning electron microscope (SEM) were used. In addition to the phylogenetic analyses of 18S rDNA, Second internal transcript spacer, 12S rDNA, cytochrome c oxidase subunit-1, and 16S rDNA were performed for molecular identification of two tick species. RESULTS: The prevalence rate of Rickettsiae in ticks was 11.6% using hemolymph staining technique and 6.17% by OmpA and gltA genes amplification. Morphological identification revealed that 100% of dogs were infested by R. sanguineus while 91.9% of camels had been infested by H. dromedarii. The phylogenetic analyses of five DNA markers confirmed morphological identification by LM and SEM. The two tick species sequences analyses proved 96-100% sequences identities when compared with the reference data in Genbank records. CONCLUSION: The present studies confirm the suitability of mitochondrial DNA markers for reliable identification of ticks at both intra- and inter-species level over the nuclear ones. In addition to, the detection of Rickettsiae in both ticks' species and establishment of the phylogenetic status of R. sanguineus and H. dromedarii would be useful in understanding the epidemiology of ticks and tick borne rickettsioses in Egypt.