Isolation and identification of multidrug resistance bacterial agents implicated in duck enteritis with first record of Salmonella enterica subspecies arizonae in Egypt.

Background: Bacterial infections causing digestive problems are among the most serious threats to Egypt's duck industry, owing to their effects on feed utilization and body weight gain. Aim: As a result, the goal of this study was to identify bacterial pathogens causing enteritis in ducks as well as testing their antimicrobials resistance capabilities. Methods: Forty-two duck flocks from different localities at four Egyptian Governorates (El-Sharkia, El-Gharbia, El-Dakahlia, and El-Qaliobia) have been subjected to clinical and postmortem examination as well as bacterial isolation and identification. The liver samples have been collected aseptically from freshly euthanized ducks for bacterial isolation followed by identification using conventional biochemical tests, VITEK 2 system, and confirmatory polymerase chain reaction (PCR) for detection of the uid A gene (beta-glucuronidase enzyme) of Escherichia coli. In addition, antimicrobial sensitivity testing for the isolates against different antimicrobials by the VITEK 2 system was used. Results: Forty-six positive bacterial isolates were identified using conventional methods and the VITEK 2 system including Staphylococcus spp. (52.17%), E. coli (41.30%), and 2.17% for each of Enterococcus casseli lavus, Salmonella enterica subspecies arizonae, and Enterobacter cloacae. PCR was positive for E. coli uid A gene at 556 bp. The antibiogram patterns of isolated pathogens from naturally infected ducks in our work demonstrated 87% multidrug resistance with varying results against different antimicrobial drugs tested. Such findings supported the fact of the upgrading multidrug resistance of Staphylococci and Enterobacteriacae. Conclusion: The most prevalent bacterial pathogens associated with duck enteritis were Staphylococcus spp. and E. coli with the first report of S. enterica subspecies arizonae causing duck enteritis in Egypt.

Therapeutic efficacy of n-Docosanol against velogenic Newcastle disease virus infection in domestic chickens.

Introduction: The control of Newcastle disease virus (NDV) infection depends solely on vaccination which in most cases is not sufficient to restrain the consequences of such a highly evolving viral disease. Finding out substances for preparing an efficient anti-ND drug would be of high value. n-Docosanol is a saturated fatty alcohol with an inhibitory effect against many enveloped viruses. In this study, we evaluated the therapeutic effect of n-docosanol on NDV infection and shedding in chickens. Methods: Chickens infected with a highly virulent NDV were treated with low to high concentrations of n-docosanol (20, 40, and 60 mg/kg body weight) for 4-successive days, once they showed the disease symptoms. Survival and curative rates, virus load, histopathological scoring, and virus shedding were defined. Results: Symptoms development was found to discontinue 24-72 hours post-treatment. Survival rate in the NDV-infected chickens raised 37.4-53.2% after the treatment. n-Docosanol treatment was also found to significantly reduce virus load in the digestive (26.2-33.9%), respiratory (38.3-63%), nervous (26.7-51.1%), and lymphatic (16.4-29.1%) tissues. Histopathological scoring of NDV lesions revealed prominent rescue effects on the histology of different tissues. Importantly, n-docosanol treatment significantly reduced virus shedding in oropharyngeal discharge and feces thereby allowing the restriction of NDV spread. Conclusion: Our findings suggest n-docosanol as a promising remedy in the control strategy of Newcastle disease in the poultry industry.

Newcastle Disease Genotype VII Prevalence in Poultry and Wild Birds in Egypt.

Newcastle Disease Virus (NDV) genotype VII is a highly pathogenic Orthoavulavirus that has caused multiple outbreaks among poultry in Egypt since 2011. This study aimed to observe the prevalence and genetic diversity of NDV prevailing in domestic and wild birds in Egyptian governorates. A total of 37 oropharyngeal swabs from wild birds and 101 swabs from domestic bird flocks including chickens, ducks, turkeys, and pelicans, were collected from different geographic regions within 13 governorates during 2019-2020. Virus isolation and propagation via embryonated eggs revealed 91 swab samples produced allantoic fluid containing haemagglutination activity, suggestive of virus presence. The use of RT-PCR targeted to the F gene successfully detected NDV in 85 samples. The geographical prevalence of NDV was isolated in 12 governorates in domestic birds, migratory, and non-migratory wild birds. Following whole genome sequencing, we assembled six NDV genome sequences (70-99% of genome coverage), including five full F gene sequences. All NDV strains carried high virulence, with phylogenetic analysis revealing that the strains belonged to class II within genotype VII.1.1. The genetically similar yet geographically distinct virulent NDV isolates in poultry and a wild bird may allude to an external role contributing to the dissemination of NDV in poultry populations across Egypt. One such contribution may be the migratory behaviour of wild birds; however further investigation must be implemented to support the findings of this study. Additionally, continued genomic surveillance in both wild birds and poultry would be necessary for monitoring NDV dissemination and genetic diversification across Egypt, with the aim of controlling the disease and protecting poultry production.

Organ-specific small non-coding RNA responses in domestic (Sudani) ducks experimentally infected with highly pathogenic avian influenza virus (H5N1).

The duck represents an important reservoir of influenza viruses for transmission to other avian and mammalian hosts, including humans. The increased pathogenicity of the recently emerging clades of highly pathogenic avian influenza (HPAI) viruses of the H5N1 subtype in ducks features systemic viral spread and organ-to-organ variation in viral transcription and tissue damage. We previously reported that experimental infection of Sudani ducks (Cairina moschata) with an Egyptian HPAI (H5N1) virus (clade 2.2.1.2) features high viral replication and severe tissue damage in lung, but lower viral replication and only mild histological changes in brain. Little is known about the involvement of miRNA in organ-specific responses to H5N1 viruses in ducks, and involvement of the other classes of small noncoding RNA (sncRNA) has not been investigated so far. Following RNA sequencing, we have annotated the duck sncRNome and compared global expression changes of the four major sncRNA classes (miRNAs, piRNAs, snoRNAs, snRNAs) between duck lung and brain during a 120 h time course of infection with this HPAI strain. We find major organ-specific differences in miRNA, piRNA and snoRNA populations even before infection and substantial reprogramming of all sncRNA classes throughout infection, which was less pronounced in brain. Pathway prediction analysis of miRNA targets revealed enrichment of inflammation-, infection- and apoptosis-related pathways in lung, but enrichment of metabolism-related pathways (including tryptophan metabolism) in brain. Thus, organ-specific differences in sncRNA responses may contribute to differences in viral replication and organ damage in ducks infected with isolates from this emerging HPAI clade, and likely other strains.

Occurrence of Salmonella genomic island 1 (SGI1) in two African Proteus mirabilis strains isolated from diseased chicken flocks.

Two P. mirabilis strains, PmSHR21 and PmSHR38, were collected from chicken flocks in Sharkia Governorate, Egypt in 2016. The two strains showed multidrug-resistance (MDR) phenotypes and were detected to harbour I) floR and sul1 genes conferring resistance to florfenicol and chloramphenicol, and sulfonamides, respectively, II) a ~1.9 kbp class 1 integron containing aadA2-lnuF genes conferring resistance to spectinomycin and streptomycin, and lincosamides, respectively. Interestingly, the two strains were detected to contain SGI1 variant, SGI1-W and inserted between the 3' end of the chromosomal trmE gene and the hipB/hipA toxin/antitoxin homologue. Fingerprinting by ERIC-PCR of the two poultry strains identified in this study and the two human SGI1-carrying P. mirabilis strains described recently in our study showed identical ERIC-pattern between SGI1-W-carrying poultry and human strains, suggesting that they might be clonally related. The detection of SGI1 and its variants in P. mirabilis isolated from humans and chicken flocks in Egypt clarify the geographical and biological spreading through an inter-transmission pathway. To the best of our knowledge, this is the first study detecting SGI1-positive P. mirabilis isolated from chicken flocks in Africa.

Evolutionary insights into the fusion protein of Newcastle disease virus isolated from vaccinated chickens in 2016 in Egypt.

Newcastle disease virus (NDV) infections are one of the most devastating causes of economic losses in the poultry industry and despite extensive vaccination, outbreaks are being reported around the globe especially from developing and tropical countries. Analysis of NDV field strains from vaccinated flocks would highlight essential areas of consideration not only to design effective immunization strategies but also to devise vaccines that provide sterile immunity. For this purpose, 91 NDV suspected outbreaks were investigated and screened for NDV genetic material. A total of 16 NDV-positive isolates were examined using biological, genetics and bioinformatics analysis to assess the epidemiological association and to identify motifs that are under vaccine-induced immune pressures. In line with the clinical outcomes, all isolates showed the 112RRQKR|F117 cleavage motif and phylogenetic analysis revealed grouping of isolates into the genotype VII, and specifically sub-genotype VIId. Further analysis of the putative fusion protein sequence showed a number of substitutions (n=10) in functionally important domains and based on these differences, the studied isolates could be categorized into four distinct groups (A-D). Importantly, two residues (N30 and K71) were conserved in the commercial live vaccine and Egyptian field strains that are present in class II, genotype II. Collectively, these data enhance our knowledge of the evolution of genotype VIId NDV under the vaccine-induced immune pressures. In addition, our findings suggest that the use of genotype II-type vaccines in Egypt may be implicated in the emergence of new variants rather than providing benefits against NDV infections.

mcr-1 identified in Avian Pathogenic Escherichia coli (APEC).

Antimicrobial resistance associated with colistin has emerged as a significant concern worldwide threatening the use of one of the most important antimicrobials for treating human disease. Here, we examined a collection (n = 980) of Avian Pathogenic Escherichia coli (APEC) isolated from poultry with colibacillosis from the US and internationally for the presence of mcr-1 and mcr-2, genes known to encode colistin resistance. Included in the analysis was an additional set of avian fecal E. coli (AFEC) (n = 220) isolates from healthy birds for comparative analysis. The mcr-1 gene was detected in a total of 12 isolates recovered from diseased production birds from China and Egypt. No mcr genes were detected in the healthy fecal isolates. The full mcr-1 gene from positive isolates was sequenced using specifically designed primers and were compared with sequences currently described in NCBI. mcr-1 positive isolates were also assessed for phenotypic colistin resistance and extended spectrum beta lactam phenotypes and genotypes. This study has identified mcr-1 in APEC isolates dating back to at least 2010 and suggests that animal husbandry practices could result in a potential source of resistance to the human food chain in countries where application of colistin in animal health is practiced.

FNR regulates expression of important virulence factors contributing to pathogenicity of uropathogenic Escherichia coli.

Uropathogenic Escherichia coli (UPEC) is responsible for the majority of urinary tract infections (UTIs), which are some of the world's most common bacterial infections of humans. Here, we examined the role of FNR (fumarate and nitrate reduction), a well-known global regulator, in the pathogenesis of UPEC infections. We constructed an fnr deletion mutant of UPEC CFT073 and compared it to the wild type for changes in virulence, adherence, invasion, and expression of key virulence factors. Compared to the wild type, the fnr mutant was highly attenuated in the mouse model of human UTI and showed severe defects in adherence to and invasion of bladder and kidney epithelial cells. Our results showed that FNR regulates motility and multiple virulence factors, including expression of type I and P fimbriae, modulation of hemolysin expression, and expression of a novel pathogenicity island involved in α-ketoglutarate metabolism under anaerobic conditions. Our results demonstrate that FNR is a key global regulator of UPEC virulence and controls expression of important virulence factors that contribute to UPEC pathogenicity.

Color image segmentation based on different color space models using automatic GrabCut.

This paper presents a comparative study using different color spaces to evaluate the performance of color image segmentation using the automatic GrabCut technique. GrabCut is considered as one of the semiautomatic image segmentation techniques, since it requires user interaction for the initialization of the segmentation process. The automation of the GrabCut technique is proposed as a modification of the original semiautomatic one in order to eliminate the user interaction. The automatic GrabCut utilizes the unsupervised Orchard and Bouman clustering technique for the initialization phase. Comparisons with the original GrabCut show the efficiency of the proposed automatic technique in terms of segmentation, quality, and accuracy. As no explicit color space is recommended for every segmentation problem, automatic GrabCut is applied with RGB, HSV, CMY, XYZ, and YUV color spaces. The comparative study and experimental results using different color images show that RGB color space is the best color space representation for the set of the images used.

Molecular and phenotypic characterization of Escherichia coli isolated from broiler chicken flocks in Egypt.

Avian pathogenic Escherichia coli (APEC) infection is responsible for great economic losses to the poultry industry worldwide and there is increasing evidence of its zoonotic importance. In this study, 219 E. coli isolates from 84 poultry flocks in Egypt, including 153 APEC, 30 avian fecal E. coli (AFEC), and 36 environmental E. coli, were subjected to phylogenetic grouping and virulence genotyping. Additionally, 50 of these isolates (30 APEC from colisepticemia and 20 AFEC) were subjected to a more-extensive characterization which included serogrouping, antimicrobial susceptibility analysis, screening for seven intestinal E. coli virulence genes (stx1, stx2, eae, espP, KatP, hlyA, and fliCh7), multilocus sequence typing (MLST), pulsed-field gel electrophoresis (PFGE), and in vivo virulence testing. More than 90% of the total APEC examined possessed iroN, ompT, hlyF, iss, and iutA, indicating that Egyptian APECs, like their counterparts from the United States, harbor plasmid pathogenicity islands (PAIs). The majority of APEC and AFEC were of phylogenetic groups A, B1, and D. For the 50-isolate subgroup, more than 70% of APEC and 80% ofAFEC were multidrug resistant. Among the subgroup of APEC, MLST analysis identified 11 sequence types (ST) while seven STs were found among AFEC. Based on PFGE, the genetic relatedness of APEC and AFEC ranged from 50%-100% and clustered into four primary groups at 50% similarity. Two of the eight APEC strains tested in chickens were able to induce 25% mortality in 1-day-old chicks. APECs were distinguished from AFECs and environmental E. coli by their content of plasmid PAI genes, whereas APEC isolated from colisepticemia and AFEC were not distinguishable based on their antimicrobial resistance patterns, as both groups were multidrug resistant. Avian E. coli strains from broiler flocks in Egypt show similar sequence types to E. coli associated with human infection.

Serum 25-hydroxyvitamin D concentration in patients with psoriasis and rheumatoid arthritis and its association with disease activity and serum tumor necrosis factor-alpha.

OBJECTIVE: To assess vitamin D status in psoriasis and rheumatoid arthritis (RA) patients and to study whether it was associated with disease activity, inflammatory markers, and serum tumor necrosis factor-alpha (TNF-α). METHODS: This cross-sectional study was conducted at Riyadh National Hospital, Riyadh, Saudi Arabia between March and September 2012. It included 43 patients with plaque psoriasis, 55 RA patients and 40 healthy controls matched for age. Blood samples were drawn from all participants for assessment of 25-hydroxyvitamin D [25(OH)D], TNF-α, C-reactive protein (CRP), erythrocyte sedimentation rate (ESR), parathyroid hormone (PTH), and serum corrected calcium. Disease activity of psoriasis and RA were assessed using Psoriasis Area and Severity Index (PASI) and Disease Activity Score Index of a 28 joint count (DAS28). RESULTS: We found a significant difference between psoriatic patients, RA patients, and healthy controls in the mean 25(OH)D (11.74±3.60, 15.45±6.42, and 24.55±11.21 ng/ml; p=0.000). We found that 25(OH)D was not correlated with PASI, DAS28, TNF-α, CRP, or ESR in psoriatic and RA patients. CONCLUSION: Serum 25-(OH)D levels are significantly lower in psoriatic and RA patients than in healthy control subjects. Low 25-OHD levels also may provide the rationale for vitamin D supplementation in the treatment of psoriasis and RA. More definitive evidence is also required to demonstrate the clinical benefit of vitamin D supplementation in the treatment of psoriasis and RA.

Proteome response of an extraintestinal pathogenic Escherichia coli strain with zoonotic potential to human and chicken sera.

A subset of extraintestinal pathogenic Escherichia coli is zoonotic and has developed strategies to adapt to different host-specific environments. However, the underlying mechanisms of these adaptive strategies have yet to be discerned. Here, the proteomic response of an avian pathogenic E. coli strain, which appears indistinguishable from neonatal meningitis E. coli, was compared following growth in human and avian sera to determine whether it uses the same mechanisms to overcome the antibacterial effects of sera from different host species. Proteins involved in biosynthesis of iron receptors were up-regulated under both sera, suggesting that serum, regardless of the host of origin, is an iron-limited environment. However, several proteins involved in synthesis of nucleic acids, sulfur-containing amino acids and fatty acids, were differentially expressed in response to the sera from different hosts. Mutational analysis showed that this APEC strain required nucleotide biosynthesis during incubation in human, but not avian serum, and deletion of genes involved in the biosynthesis of sulfur-containing amino acids increased its resistance to human serum. Continued investigation of the proteome of 'zoonotic' ExPEC strains, grown under other 'dual' host conditions, will contribute to our understanding of ExPEC pathogenesis and host specificity and development of effective therapies and control strategies.

Avian-pathogenic Escherichia coli strains are similar to neonatal meningitis E. coli strains and are able to cause meningitis in the rat model of human disease.

Escherichia coli strains causing avian colibacillosis and human neonatal meningitis, urinary tract infections, and septicemia are collectively known as extraintestinal pathogenic E. coli (ExPEC). Characterization of ExPEC strains using various typing techniques has shown that they harbor many similarities, despite their isolation from different host species, leading to the hypothesis that ExPEC may have zoonotic potential. The present study examined a subset of ExPEC strains: neonatal meningitis E. coli (NMEC) strains and avian-pathogenic E. coli (APEC) strains belonging to the O18 serogroup. The study found that they were not easily differentiated on the basis of multilocus sequence typing, phylogenetic typing, or carriage of large virulence plasmids. Among the APEC strains examined, one strain was found to be an outlier, based on the results of these typing methods, and demonstrated reduced virulence in murine and avian pathogenicity models. Some of the APEC strains tested in a rat model of human neonatal meningitis were able to cause meningitis, demonstrating APEC's ability to cause disease in mammals, lending support to the hypothesis that APEC strains have zoonotic potential. In addition, some NMEC strains were able to cause avian colisepticemia, providing further support for this hypothesis. However, not all of the NMEC and APEC strains tested were able to cause disease in avian and murine hosts, despite the apparent similarities in their known virulence attributes. Thus, it appears that a subset of NMEC and APEC strains harbors zoonotic potential, while other strains do not, suggesting that unknown mechanisms underlie host specificity in some ExPEC strains.