Lucrative antioxidant effect of metformin against cyclophosphamide induced nephrotoxicity.

Cyclophosphamide is anticancer drug with a well-Known nephrotoxicity. This work was applied to study the lucrative antioxidant influence of metformin as co-therapy on the nephrotoxicity induced by cyclophosphamide in the treatment of different cancer diseases. Four groups of male Sprague Dawley rats were used; Control group (C) received single I.P. injection of 0.2 ml saline, Metformin (MET) group received daily gavage of 200 mg/kg metformin for two weeks, Cyclophosphamide (CP) group received single I.P. injection of 200 mg/kg CP, Protector group (CP.MET) received daily gavage of 200 mg/kg metformin for two weeks and single I.P. injection of 200 mg/kg CP at day 7. By day 14 rats were euthanized. Samples were collected from kidney tissues and blood for kidney function evaluation, histopathological and assessment of oxidative stress markers. The results disclosed that CP yields many functional and structural damage to the kidney, worsened oxidative stress markers and kidney function indicators. The protector group displayed better kidney tissue morphology, acceptable kidney function indicators as well as satisfactory oxidative stress markers. In assumption, metformin could be combined with CP owing to its lucrative effect counter to CP persuaded nephrotoxicity.

Multidrug-resistant Escherichia coli in Raw Milk: Molecular Characterization and the potential impact of camel's Urine as an Antibacterial Agent.

Raw milk is one of the most important vehicles for transmitting various pathogens, especially Escherichia coli (E. coli). Multidrug-resistant pathogens are highly prevalent among mastitic cows in various dairy farms worldwide. Therefore, our current study is based on the identification of E. coli from mastitic cow's milk and their resistance to various antibacterial agents. As well, the impact of camel's urine on multi-drug resistant E. coli were also evaluated. Thirty-three E. coli isolates were recovered from 254 milk samples. All strains were initially identified phenotypically by culturing on specific media and Vitek 2 Compact System. The protein fingerprinting technique was used as a confirmatory method. The Stx1, Stx2 and eae genes were also verified by polymerase chain reaction (PCR). The antimicrobial resistance of E. coli strains was tested by the Vitek 2 AST-GN69 cards. Thirty multi-drug resistant E. coli strains (20 from mastitic milk and 10 from clinical samples) were laboratory tested with different concentrations (100%, 75%, 50% and 25%) of virgin and breeding camel's urine, using the paper disc diffusion method. Our findings showed that 93.94% of E. coli strains were recognized by the Vitek™ 2 system. The results of proteomic investigation illustrated that 100% of E. coli strains were identified at log values ≥2.00. The genotypic identification of the three virulence genes illustrated that 90.1%, 63.64%, and 30.55% of E. coli strains were able to carry the Stx1, eae, and Stx2 genes, respectively. Most strains of E. coli showed strong resistance against cefazolin (78.79%), ceftazidime (66.67%), cefotaxime (60.61%), ceftriaxone (54.55%), and cefepime (39.40%). The results of the antibacterial effect of camel's urine revealed that the mean inhibitory zones of virgin camel's urine were 28 mm, 17 mm, and 14 mm, for the concentrations of 100%, 75%, and 50%, respectively. Whereas; the inhibitory zones for the breeding camel's urine were 18 mm, 0 mm, and 0 mm, for the concentrations of 100%, 75%, and 50%, respectively. We concluded that the majority of E. coli strains were able to harbor some virulence genes and resist many antibiotics. Our study also provided a robust evidence that the camel's urine, particularly from the virgin camels has robust antimicrobial activity against multidrug-resistant E. coli strains.

Antibiogram and phylogenetic diversity of enterotoxigenic Staphylococcus aureus strains from milk products and public health implications.

Food poisoning caused by Staphylococcus aureus (S. aureus) toxins is considered one of the foremost public health threat that usually occurs through the ingestion of raw milk contaminated with staphylococcal enterotoxins. The current study spotlights on the prevalence, antibiogram and genetic diversity of S. aureus enterotoxin genes. One hundred and fifty of raw milk (90) and ice cream (60) samples were randomly collected from local markets from Sadat city, Egypt. S. aureus was recovered from 44% of raw milk and 20% of ice cream samples. The identification for the obtained S. aureus isolates was confirmed through targeting the nuc gene. Antibiogram pattern of 32 S. aureus isolates showed high resistance to Cefoxitin, Sulpha/Trimethoprim, Tetracycline, Norfloxacin, Penicillin and Cephradine. However, high susceptibility to Gentamycin and Vancomycin were observed. Multiplex PCR was a competent practise for the recognition of Staphylococcus enterotoxin (SE) genes (SEA, SEB and SED). The phylogenetic analysis of the SED gene of enterotoxigenic S. aureus strains showed identical similarity with 100% to each other and high similarity with other international isolates in GenBank from different localities and sources. The frequency of enterotoxigenic S. aureus strains in milk products could have serious hazardous effects on humans. These results suggested possible strains transmission between different geographical areas through the food and milk product trades.

A novel bivalent Pasteurellosis-RHD vaccine candidate adjuvanted with Montanide ISA70 protects rabbits from lethal challenge.

In the present study, a bivalent vaccine against Pasteurella multocida and rabbit hemorrhagic disease virus (RHDV) was formulated with Montanide™ ISA70 oil adjuvant (Seppic, Paris, France). Its efficacy was evaluated and compared to similar monovalent preparations and commercially available monovalent vaccines. White new Zeeland rabbit groups (n = 10) received 2 successive doses of the tested vaccines and were challenged 2 weeks after 2nd dose with Pasteurella multocida and RHDV or either pathogens according to their vaccination schedule. Challenged not-vaccinated group of rabbits (n = 10) was included as a control. The bivalent and monovalent ISA70 preparations were found stable, safe, sterile, pure and of low viscosity. Group 3 (GP3) which received bivalent vaccine showed the highest antibody geometric mean titers against Pasteurella multocida and RHDV evaluated by ELISA and hemagglutination inhibition (HI) respectively. Following virulent challenge; Gp3 rabbits were 90% protected from challenge over other groups that showed 80% protection. Detection of either pathogen in the livers of dead and euthanized rabbits had failed except for non-vaccinated controls. The bivalent vaccine candidate was fully protective. Immunization against both pathogens can be achieved by single vaccination.

Genetic diversity of Salmonella enterica recovered from chickens farms and its potential transmission to human.

BACKGROUND: Salmonella is a zoonotic bacterium transmitted through the food chain and is an important cause of disease in humans. The current study is aimed to characterize Salmonella isolates from broiler breeder chickens farms using, polymerase chain reaction (PCR) and sequencing analysis of representative isolates. METHODS: S. Kentucky (n=11), S. Enteritidis (n=4), S. Typhimurium (n=3), S. Breanderp (n=1), and Sand S. Newport (n=1), were identified from chicken farms. Antimicrobial sensitivity test among the strains were investigated using 13 antibacterial discs. The amplified fragments of fliC and sefA genes were used to characterize S. Kentucky, S. Enteritidis and S. Typhimurium strains. Sequence analysis of the amplified PCR products for Salmonella Kentucky, Enteritidis and Typhimurium were carried out. RESULTS: Antimicrobial sensitivity testing revealed that 95% of the isolates were resistant to penicillin, 85% to norfloxacin and colistin sulfate (each), 75% to gentamicin, 70% to nalidixic acid and 60% to flumequine. The obtained sequences revealed the close identity of the isolated strains with other Salmonella reference strains in different countries. CONCLUSION: Analysis of the selected salmonellae confirm the report of Salmonella Enteritidis, Salmonella Typhimurium and Salmonella Kentucky circulation among broiler breeder flocks and the need to determine antibacterial susceptibility pattern regularly to detect multidrug-resistant salmonellae. The present study reports the circulation of Salmonella Kentucky, Enteritidis and Typhimurium among broiler breeder farms in Egypt. Emergency control of salmonellae is a global public health concern.

Alkaline serine protease from the new halotolerant alkaliphilic Salipaludibacillus agaradhaerens strain AK-R: purification and properties.

Herein, we report the purification and characterization of an alkaline protease from the alkaliphilic Salipaludibacillus agaradhaerens (formerly Bacillus agaradhaerens) strain AK-R, which was previously isolated from Egyptian soda lakes. The purification procedures resulted in enzyme purification up to 13.3-fold, with a recovery yield of 16.3% and a specific activity of 3488 U/mg protein. AK-R protease was a monomeric protein with an estimated molecular weight of 33.0 kDa. The optimum pH and temperature for AK-R protease were pH 10 and 60 °C, respectively. The enzyme thermostability was significantly enhanced in the presence of CaCl2 by approximately 1.3-fold. Moreover, under optimal conditions, the K m and V max values of the enzyme were 2.63 mg/ml and 4166.7 U/mg, respectively. PMSF caused complete inhibition of the enzyme activity, suggesting that AK-R belongs to the serine protease family. In addition, the enzyme was completely inhibited by EDTA, revealing the requirement of metal ions for AK-R protease activity; hence, it can be classified as a metalloprotease. AK-R protease is a mostly thiol-independent enzyme, since thiol reductants such as ß-mercaptoethanol and dithiothreitol had no effect on the enzyme activity. AK-R protease exhibited high stability in several organic solvents, including butanol, amyl alcohol, dimethyl ether, toluene, diethyl ether and methanol. Moreover, AK-R protease showed significant stability to a variety of surfactants and commercial detergents. The features and properties of AK-R alkaline protease are favourable and suggest its potential applications in various industries, particularly in the laundry detergent industry.

Occurrence of Salmonella infection and antimicrobial susceptibility for local Salmonella isolates from different sources in a cross-sectional study.

Salmonellosis is a considerable public health problem worldwide, with high economic importance in developed countries. The main purpose of this study was to determine the prevalence of Salmonella infection and antibiogram analysis of isolated strains in a cross-sectional study in Egypt 2016-2017. The study investigated twenty-eight Salmonella isolates from different areas in Egypt and different types of samples, such as human stool (9.3%), Egyptian cattle egrets and storks (28.5%) and grilled chicken from electric grills (36.6%). No isolates were detected from grilled chicken from charcoal grills or drinking water. The main Salmonella serotype detected in the isolates was S. typhimurium (86.5%). Molecular characterization of the invA gene by PCR was carried out and then confirmed by sequencing, and the results were submitted to GenBank. Antibiogram analysis of Egyptian isolates carried out on 9 antimicrobial discs reported that the routine regimes of treatment were not yet effective for recent new Salmonella generations in 2016-2017. The new isolates could be treated with levofloxacin, cefaperazone/sulbactam, chloramphenicol, imipenem or meropenem.

Poultry and beef meat as potential seedbeds for antimicrobial resistant enterotoxigenic Bacillus species: a materializing epidemiological and potential severe health hazard.

Although Bacillus cereus is of particular concern in food safety and public health, the role of other Bacillus species was overlooked. Therefore, we investigated the presence of eight enterotoxigenic genes, a hemolytic gene and phenotypic antibiotic resistance profiles of Bacillus species in retail meat samples. From 255 samples, 124 Bacillus isolates were recovered, 27 belonged to B. cereus and 97 were non-B. cereus species. Interestingly, the non-B. cereus isolates carried the virulence genes and exhibited phenotypic virulence characteristics as the B. cereus. However, correlation matrix analysis revealed the B. cereus group positively correlates with the presence of the genes hblA, hblC, and plc, and the detection of hemolysis (p < 0.05), while the other Bacillus sp. groups are negatively correlated. Tests for antimicrobial resistance against ten antibiotics revealed extensive drug and multi-drug resistant isolates. Statistical analyses didn't support a correlation of antibiotic resistance to tested virulence factors suggesting independence of these phenotypic markers and virulence genes. Of special interest was the isolation of Paenibacillus alvei and Geobacillus stearothermophilus from the imported meat samples being the first recorded. The isolation of non-B. cereus species carrying enterotoxigenic genes in meat within Egypt, suggests their impact on food safety and public health and should therefore not be minimised, posing an area that requires further research.

Characterization and susceptibility of streptococci and enterococci isolated from Nile tilapia (Oreochromis niloticus) showing septicaemia in aquaculture and wild sites in Egypt.

BACKGROUND: The present investigation was an endeavor into the elucidation of the disease-causing pathogen of streptococcosis in Nile tilapia (Oreochromis niloticus) in Egypt affecting adult fish cultured and wild fish in the Nile river. Fish were obtained from commercial fishermen, collected as part of their routine fishing activities. The researchers observed the routine fishing process and selected fish for use in the study, at the point of purchase from the fisherman. RESULTS: Diseased fish showed exophthalmia with accumulation of purulent and haemorrhagic fluid around eyes, and ventral petechial haemorrhages. The Post mortem examination revealed, abdominal fat haemorrhage, pericarditis and enlargement of the liver, spleen and kidney. Gram-stained smears revealed the presence of Gram-positive cocci, ß-hemolytic, oxidase and catalase negative. Analysis of the 16S rRNA gene confirmed that the 17 tilapia isolates studied were 6/17 Enterococcus faecalis, 2/17 Enterococcus gallinarum, 3/17 Streptococcus pluranimalium, 2/17 Aerococcus viridans, 1/17 isolate of each Streptococcus dysgalactiae, Streptococcus anginosus, Lactococcus garvieae and Granulicetella elegans/Leuconostoc mesenteroides cremoris. It should be noted that there was no mixed infection. Multiple resistance was observed and the most frequent antibiotic combination was penicillin, ampicillin, vancomycin, chloramphenicol, rifampicin, ofloxacin, clindamycin, erythromycin and tetracycline representing eight classes. CONCLUSIONS: Consequently, we concluded that Streptococcus species are an emerging pathogen for Nile tilapia aquaculture in Egypt and to be considered as a new candidate in the warm water fish diseases in Egypt with special reference to L. garvieae, S. dysgalactiae in addition to L. mesenteroides cremoris which was not reported before from tilapia and taking into consideration their zoonotic implications for public health.

Characterization and anti-Aspergillus flavus impact of nanoparticles synthesized by Penicillium citrinum.

This work was conducted to evaluate the ability of grape molding fungus; Penicillium citrinum to synthesize silver nanoparticles (Ag NPs). The potency of biosynthesized Ag NPs was checked against the aflatoxigenic Aspergillus flavus var. columnaris, isolated from sorghum grains. Biosynthesized Ag NPs were characterized and confirmed in different ways. X ray diffraction (XRD), Energy Dispersive Spectroscopy (EDS), Transmission Electron Microscopy (TEM) and optical absorption measurements confirmed the bio-synthesis of Ag NPs. The in vitro antifungal investigation showed that biosynthesized Ag NPs were capable of inhibiting the growth of aflatoxigenic A. flavus var. columnaris. Utilization of plant pathogenic fungi in the Ag NPs biosynthesis as well as the use of bio-Ag NPs to control fungal plant diseases instead of chemicals is promising. Further work is needed to confirm the efficacy of the bio-Ag NPs against different mycotoxigenic fungi and to determine the potent applicable doses.

Antimicrobial resistance and virulence characterization of Staphylococcus aureus and coagulase-negative staphylococci from imported beef meat.

BACKGROUND: The objectives of this study were to characterize the diversity and magnitude of antimicrobial resistance among Staphylococcus species recovered from imported beef meat sold in the Egyptian market and the potential mechanisms underlying the antimicrobial resistance phenotypes including harboring of resistance genes (mecA, cfr, gyrA, gyrB, and grlA) and biofilm formation. RESULTS: The resistance gene mecA was detected in 50% of methicillin-resistant non-Staphylococcus aureus isolates (4/8). Interestingly, our results showed that: (i) resistance genes mecA, gyrA, gyrB, grlA, and cfr were absent in Staphylococcus hominis and Staphylococcus hemolyticus isolates, although S. hominis was phenotypically resistant to methicillin (MR-non-S. aureus) while S. hemolyticus was resistant to vancomycin only; (ii) S. aureus isolates did not carry the mecA gene (100%) and were phenotypically characterized as methicillin- susceptible S. aureus (MSS); and (iii) the resistance gene mecA was present in one isolate (1/3) of Staphylococcus lugdunensis that was phenotypically characterized as methicillin-susceptible non-S. aureus (MSNSA). CONCLUSIONS: Our findings highlight the potential risk for consumers, in the absence of actionable risk management information systems, of imported foods and advice a strict implementation of international standards by different venues such as CODEX to avoid the increase in prevalence of coagulase positive and coagulase negative Staphylococcus isolates and their antibiotic resistance genes in imported beef meat at the Egyptian market.

Molecular characterization of the capsular antigens of Pasteurella multocida isolates using multiplex PCR.

The use of molecular techniques for detection and characterization of the Pasteurella multocida is very important for rapid and specific detection and characterization of the organism. During the period from 15th February, 2014 to 15th April, 2015, 425 nasopharyngeal swabs and 175 lung and spleen samples were collected and examined by conventional methods, 80 strains (18.82%) of P. multocida were isolated from the calves, sheep and goat with respiratory manifestation. Meanwhile, 77 strains (44%) were isolated from emergency slaughtered animals. All the recovered strains were positive for specific PCR for detection of P. multocida strains previously identified as P. multocida by standard microbiological techniques. Multiplex PCR for molecular typing of the capsular antigens of the recovered P. multocida revealed positive amplification of 1044 bp fragments specific to the capsular antigen type A with 105 strains (66.88%), and amplification 511 bp fragments of the capsular antigen type E with 52 strain (33.12%) and absence of B, D and F antigens. Multiplex PCR for molecular typing of the capsular antigens of P. multocida can be used as a simple, sensitive, rapid, reliable technique instead of the serological techniques for identification of the capsular antigens of P. multocida.

Prevalence of the Antibiotic Resistance Genes in Coagulase-Positive-and Negative-Staphylococcus in Chicken Meat Retailed to Consumers.

The use of antibiotics in farm management (growing crops and raising animals) has become a major area of concern. Its implications is the consequent emergence of antibiotic resistant bacteria (ARB) and accordingly their access into the human food chain with passage of antibiotic resistance genes (ARG) to the normal human intestinal microbiota and hence to other pathogenic bacteria causative human disease. Therefore, we pursued in this study to unravel the frequency and the quinolone resistance determining region, mecA and cfr genes of methicillin-susceptible Staphylococcus aureus (MSSA), methicillin-resistant S. aureus (MRSA), methicillin-resistant coagulase-negative staphylococci (MRCNS) and methicillin-susceptible coagulase-negative staphylococci (MSCNS) isolated from the retail trade of ready-to-eat raw chicken meat samples collected during 1 year and sold across the Great Cairo area. The 50 Staphylococcus isolated from retail raw chicken meat were analyzed for their antibiotic resistance phenotypic profile on 12 antibiotics (penicillin, oxacillin, methicillin, ampicillin-sulbactam, erythromycin, tetracycline, clindamycin, gentamicin, ciprofloxacin, chloramphenicol, sulfamethoxazole-trimethoprim, and vancomycin) and their endorsement of the quinolone resistance determining region, mecA and cfr genes. The isolation results revealed 50 isolates, CPS (14) and CNS (36), representing ten species (S. aureus, S. hyicus, S. epidermedius, S. lugdunensis, S. haemolyticus, S. hominus, S. schleiferi, S. cohnii, S. intermedius, and S. lentus). Twenty seven isolates were methicillin-resistant. Out of the characterized 50 staphylococcal isolates, three were MRSA but only 2/3 carried the mecA gene. The ARG that bestows resistance to quinolones, ß-lactams, macrolides, lincosamides, and streptogramin B [MLS(B)] in MRSA and MR-CNS were perceived. According to the available literature, the present investigation was a unique endeavor into the identification of the quinolone-resistance-determining-regions, the identification of MRSA and MR-CNS from retail chicken meat in Egypt. In addition, these isolates might indicate the promulgation of methicillin, oxacillin and vancomycin resistance in the community and imply food safety hazards.

Phenotypic and genotypic analysis of pathogenic Escherichia coli virulence genes recovered from Riyadh, Saudi Arabia.

The current study was carried out to evaluate the phenotypic and genotypic characterization of avian pathogenic Escherichia coli recovered from Riyadh, Saudi Arabia. During the period of 10th February-30th May 2015, 70 E. coli strains were isolated from chicken farms located in Riyadh, Saudi Arabia. All strains were tested phenotypically by standard microbiological techniques, serotyped and the virulence genes of such strains were detected by polymerase chain reaction (PCR). Most of the recovered strains from chickens belonged to serotype O111:K58 25 strains (35.7%), followed by serotype O157:H7 13 strains (18.57%), followed by serotype O114:K90 10 strains (14.29%), then serotype O126:K71 9 strains (12.9%), serotype O78:K80 8 strains (11.43%) and in lower percentage serotype O114:K90 and O119:K69 5 strains (7.14%). The virulence genotyping of E. coli isolates recovered from broilers revealed the presence of the uidA gene in all the field isolates (6 serovars) examined in an incidence of 100%, as well as the cvaC gene was also present in all field isolates (6 serovars), while the iutA gene and the iss gene were detected in 5 out of 6 field serovars in an incidence of 81.43% and 64.29%, respectively. Phenotypical examination of the other virulence factors revealed that 65 isolates were hemolytic (92.9%), as well as 15 isolates (21.42%) were positive for enterotoxin production. Meanwhile, 21 isolates (30%) were positive for verotoxin production, 58 isolates (82.86%) for the invasiveness and 31 isolates (44.29%) for Congo red binding activities of the examined serotypes.