Phenotypic and Genotypic Screening of Colistin Resistance Associated with Emerging Pathogenic Escherichia coli Isolated from Poultry.

Chickens continue to be an important reservoir of zoonotic multidrug-resistant illnesses. Antimicrobial resistance correlated with colistin has emerged as a critical concern worldwide in the veterinary field and the public health sector. The current study investigated the prevalence of multidrug-resistant avian pathogenic Escherichia coli among chicken farms in three Egyptian governorates, focusing on colistin resistance assessment. A total of 56 Escherichia coli isolates were recovered out of 120 pooled samples obtained from diseased chicken broilers (46.7%). The E. coli isolates were serotyped to nine different serotypes; the highest incidence was for O125 (n = 18). The E. coli isolates demonstrated multidrug-resistant patterns against 10 antibiotics, especially clindamycin, tetracycline, streptomycin and ampicillin, by 100, 100, 96.4 and 92.9%, respectively. On the other hand, colistin resistance was 41.1% using AST. All E. coli isolates displayed positive colistin resistance growth on chromogenic medium, but only 25% represented this positivity via MIC estimation and Sensititre kit. PCR results revealed that all isolates harbored mcr-1, but no isolates harbored the other 2-5 mcr genes. In conclusion, the study demonstrated the emergence of multidrug-resistant, especially colistin-resistant, E. coli among chicken broiler flocks, and mcr-1 is the master gene of the colistin resistance feature.

Incorporation of Plant Extracted Hydroxyapatite and Chitosan Nanoparticles on the Surface of Orthodontic Micro-Implants: An In-Vitro Antibacterial Study.

In our study, the structural and morphological applications of hydroxyapatite and chitosan nanoparticles and coated micro-implants were assessed for their ability to combat oral pathogenic bacteria. The hydroxyapatite, as well as chitosan nanoparticles, were synthesized from the Salvadora persica plant. The crystal morphology, phase composition, particle size, and surface functional groups of the nano-samples were analyzed via classical examinations and energy dispersive X-ray analysis. The prepared nanoparticles have been examined for antibacterial activity against four common oral bacterial strains. The antimicrobial effect was also assessed by the Live/Dead BacLight technique in combination with confocal scanning laser microscopy. Titanium micro-implants were coated with regular hydroxyapatite (HAP) and chitosan nanoparticles, and the surface was characterized by scanning electron microscopy. The analysis asserted elemental composition of the prepared nanoparticles and their textural features, metal crystallization, and functional bonds. The antibacterial activity of the nanoparticles was evaluated against oral pathogenic microorganisms by the disc diffusion method, minimum bacterial concentration (MBC), and minimum inhibitory concentration (MIC). Chitosan nanoparticles showed (MICs) of 8 µg mL-1 for (Streptococcus salivarius, Streptococcus mutans and Enterococcus faecalis), and 16 µg mL-1 for Streptococcus sanguinis. HAP nanoparticles showed (MICs) of 16 µg/mL for E. faecalis, and S. sanguis, 8 µg/mL for S. salivarius and finally 4 µg/mL for S. mutans. HAP nanoparticles showed enhanced antibacterial activity and more obvious damage in the bacterial cell membrane than that of synthesized chitosan nanoparticles. The prepared nanoparticles could successfully coat titanium microplates to enhance their efficiency.

Advanced molecular characterization of enteropathogenic Escherichia coli isolated from diarrheic camel neonates in Egypt.

BACKGROUND AND AIM: Camels are important livestock in Egypt on cultural and economic bases, but studies of etiological agents of camelid diseases are limited. The enteropathogen Escherichia coli is a cause of broad spectrum gastrointestinal infections among humans and animals, especially in developing countries. Severe infections can lead to death. The current study aimed to identify pathogenic E. coli strains that cause diarrhea in camel calves and characterize their virulence and drug resistance at a molecular level. MATERIALS AND METHODS: Seventy fecal samples were collected from diarrheic neonatal camel calves in Giza Governorate during 2018-2019. Samples were cultured on a selective medium for E. coli, and positive colonies were confirmed biochemically, serotyped, and tested for antibiotic susceptibility. E. coli isolates were further confirmed through detection of the housekeeping gene, yaiO, and examined for the presence of virulence genes; traT and fimH and for genes responsible for antibiotic resistance, ampC, aadB, and mphA. The isolates in the important isolated serotype, E. coli O26, were examined for toxigenic genes and sequenced. RESULTS: The bacteriological and biochemical examination identified 12 E. coli isolates from 70 fecal samples (17.1%). Serotyping of these isolates showed four types: O26, four isolates, 33.3%; O103, O111, three isolates each, 25%; and O45, two isolates, 16.7%. The isolates showed resistance to vancomycin (75%) and ampicillin (66.6%), but were highly susceptible to ciprofloxacin, norfloxacin, and tetracycline (100%). The structural gene, yaiO (115 bp), was amplified from all 12 E. coli isolates and traT and fimH genes were amplified from 10 and 8 isolates, respectively. Antibiotic resistance genes, ampC, mphA, and aadB, were harbored in 9 (75%), 8 (66.6%), and 5 (41.7%), respectively. Seven isolates (58.3%) were MDR. Real-time-polymerase chain reaction of the O26 isolates identified one isolate harboring vt1, two with vt2, and one isolate with neither gene. Sequencing of the isolates revealed similarities to E. coli O157 strains. CONCLUSION: Camels and other livestock suffer various diseases, including diarrhea often caused by microbial pathogens. Enteropathogenic E. coli serotypes were isolated from diarrheic neonatal camel calves. These isolates exhibited virulence and multiple drug resistance genes.

Multidrug-resistant strains of Mycobacterium complex species in Egyptian farm animals, veterinarians, and farm and abattoir workers.

BACKGROUND AND AIM: Mycobacterium tuberculosis complex (MTBC) is a group of mycobacteria that are important human pathogens. Mycobacterium tuberculosis and Mycobacterium bovis cause serious chronic life-threatening disease and also significant economic losses in both production and remedication. Recently, emergence of multidrug-resistant tuberculosis (MDR-TB) complex has generated global recognition of the need for rapid and sensitive diagnosis and development of new treatments. The current study illustrates the isolation/identification of MTBC strains in specimens obtained from cows and humans by conventional and real-time polymerase chain reaction (RT-PCR) techniques. Further, the study assesses sensitivity to antituberculosis drugs in isolated MDR strains. MATERIALS AND METHODS: A total of 1464 samples from cattle (1285 raw milk and 179 lymph node), and 149 human sputum samples, were collected from farms and abattoirs in Delta Egypt. Conventional methods (culture and Ziehl-Neelsen staining) were implemented as were RT-PCR using MTBC universal DNA. The effect of some antituberculosis drugs on obtained isolates was assayed using drug susceptibility proportion and qualitative suspension techniques. RESULTS: The MBTC detection rate using the culture method was higher than for Ziehl-Neelsen staining; raw cow milk (2.56 vs. 1.63%), lymph nodes (51.59 vs. 48.04%), and human sputum (5.36 vs. 4.02%). A total of 135 isolates were obtained. Application of RT-PCR detected 138 isolates from the same set of samples. MBTC isolates were resistant to first-line antituberculosis drugs, such as pyrazinamide, isoniazid, rifampicin, and ethambutol by 78.5, 59.3, 40.7, and 31.8%, respectively, and could be highly resistant to kanamycin (82.3%) and amikacin (80.7%). However, isolates remained sensitive to ciprofloxacin (71.1%) and clarithromycin (73.3%) as second-line drugs. CONCLUSION: There is a growing risk for isolation of MDR-TB from raw milk and lymph nodes of field tuberculin positive cattle as well as sputum of veterinarians and workers existed in farms and abattoirs. PCR-based techniques have become the gold standard for the identification of mycobacterial species, showing high efficiency compared to bacteriological and microscopic examination. Application of the first- and second-line antituberculosis drugs in combination could counter the MDR-TB concern once infections are identified.

Evaluation of specific and non-specific immune response of four vaccines for caseous lymphadenitis in sheep challenged.

BACKGROUND: Caseous lymphadenitis (CLA) is a serious disease affects sheep and goat, caused by Corynebacterium pseudotuberculosis. Due to it is non-treatable disease, so the effective preventive vaccines are considered a significant way to combat the disease. All strains of C. pseudotuberculosis have several virulence factors that associated with their cell invasion, survival, and proliferation such as phospholipase D (PLD), outer lipid coat, and secreted proteases. AIM: The present study was directed to perform a comparative innate and acquired immune response assessment of different four vaccine formulas to evoke protection against induced (CLA) challenge in sheep. MATERIALS AND METHODS: Negative ELISA (free of CLA) 15 local breed male (Balady) sheep were divided into five groups, each has received a different vaccine while the control has received saline buffer. The first vaccine composed of toxoid PLD alone the second composed of toxoid PLD with bacterin (formalinkilled bacteria), the third vaccine composed of toxoid PLD plus covaccine 8, while the fourth one composed of toxoid PLD plus locally produced polyvalent clostridial vaccine. The specific immune response was evaluated through lymphocyte proliferation assay using ELISA BrdU kit, while the non-specific response was estimated by superoxide anion production and lysozyme activity assays. RESULTS: The study revealed that PLD toxoid could evoke the highest specific immune response, showing a stimulation index (9.12%). On the other hand, combined toxoid ↱PLD with bacterin followed by PLD toxoid showed a significant increase in the non-specific innate immune response. CONCLUSION: The present study indicated that the toxoid PLD alone vaccine was most efficient and provided innate and acquired immune response in animals against CLA.

Serotyping, antibiotic susceptibility, and virulence genes screening of Escherichia coli isolates obtained from diarrheic buffalo calves in Egyptian farms.

AIM: In Egypt as in many other countries, river water buffalo (Bubalus bubalis) is considered an important source of high-quality milk and meat supply. The objective of this study was to investigate serotypes, virulence genes, and antibiotic resistance determinants profiles of Escherichia coli isolated from buffalo at some places in Egypt; noticibly, this issue was not discussed in the country yet. MATERIALS AND METHODS: A number of 58 rectal samples were collected from diarrheic buffalo calves in different regions in Egypt, and bacteriological investigated for E. coli existence. The E. coli isolates were biochemically, serologicaly identified, tested for antibiotic susceptibility, and polymerase chain reaction (PCR) analyzed for the presence of antibiotic resistance determinants and virulence genes. RESULTS: Overall 14 isolates typed as E. coli (24.1%); 6 were belonged to serogroup O78 (10.3%), followed by O125 (4 isolates, 6.9%), then O158 (3 isolates, 5.2%) and one isolate O8 (1.7%), among them, there were 5 E. coli isolates showed a picture of hemolysis (35.7%). The isolates exhibited a high resistance to ß lactams over 60%, followed by sulfa (50%) and aminoglucoside (42.8%) group, in the same time the isolates were sensitive to quinolone, trimethoprim-sulfamethoxazole, tetracycline (100%), and cephalosporine groups (71.4%). A multiplex PCR was applied to the 14 E. coli isolates revealed that all were carrying at least one gene, as 10 carried blaTEM (71.4%), 8 Sul1 (57.1%), and 6 aadB (42.8%), and 9 isolates could be considered multidrug resistant (MDR) by an incidence of 64.3%. A PCR survey was stratified for the most important E. coli virulence genes, and showed the presence of Shiga toxins in 9 isolates carried either one or the two Stx genes (64.3%), 5 isolates carried hylA gene (35.7%), and eae in 2 isolates only (14.3%), all isolates carried at least one virulence gene except two (85.7%). CONCLUSION: The obtained data displayed that in Egypt, buffalo as well as other ruminants could be a potential source of MDR pathogenic E. coli variants which have a public health importance.