Antibacterial and Anti-Biofilm Activities of Cinnamon Oil against Multidrug-Resistant Klebsiella pneumoniae Isolated from Pneumonic Sheep and Goats.

The primary objectives were to isolate and identify Klebsiella pneumoniae (K. pneumoniae), and determine the antimicrobial resistance patterns and biofilm formation abilities of the isolates. Additionally, the study aimed to investigate the antimicrobial and anti-biofilm effects of cinnamon oil against K. pneumoniae isolates. A cross-sectional study was conducted from March 2022 to April 2023 to collect 200 samples (including 156 nasal swabs and 44 lung specimens) from pneumonic sheep and goats admitted to the Veterinary Teaching Hospital of Zagazig University, Egypt. K. pneumoniae was isolated from a total of 72 (36%) samples, with 53 (73.6%) isolates recovered from nasal swabs and 19 (26.4%) from lung samples. Among the samples, 52 (36.9%) were from sheep and 20 (33.9%) were from goats. Antimicrobial susceptibility testing of the 72 K. pneumoniae isolates to 18 antimicrobials revealed that all isolates were resistant to ampicillin, amoxicillin/clavulanic acid, cefotaxime, ceftriaxone, tetracycline, colistin, fosfomycin, and trimethoprim/sulphamethoxazole. None of the isolates were resistant to amikacin, imipenem, and norfloxacin. Multidrug resistance (MDR) was observed in all K. pneumoniae isolates recovered from sheep and goats. The average MAR index was 0.71, ranging from 0.50 to 0.83. Regarding biofilm formation, among the K. pneumoniae isolates with a high MAR index (n = 30), 10% exhibited strong formation, 40% showed moderate formation, 43.3% displayed weak formation, and 6.7% did not form biofilms. Additionally, the biofilm-forming genes treC and fimA were present in all 28 biofilm-forming K. pneumoniae isolates, while the mrkA gene was detected in 15 (53.6%) of the 28 isolates. MDR K. pneumoniae isolates with strong biofilm formation abilities were treated with cinnamon oil at varying concentrations (100%, 75%, 50%, and 25%). This treatment resulted in inhibition zone diameters ranging from 35 to 45 mm. Cinnamon oil exhibited lower minimum inhibitory concentration and minimum bactericidal concentration values compared to norfloxacin for all isolates. Additionally, cinnamon oil significantly reduced the expression of biofilm-associated genes (treC, fimA, and mrkA) when compared to isolates treated with norfloxacin or untreated. In conclusion, this study identified a high level of MDR K. pneumoniae with strong and moderate biofilm formation abilities in pneumonic sheep and goats in Sharika Governorate, Egypt. Although cinnamon oil demonstrated potential antibacterial and anti-biofilm properties against K. pneumoniae, further research is required to investigate its effectiveness in treating K. pneumoniae infections in pneumonic sheep and goats.

Molecular Characterization and Antimicrobial Susceptibilities of Corynebacterium pseudotuberculosis Isolated from Caseous Lymphadenitis of Smallholder Sheep and Goats.

Caseous lymphadenitis (CLA) is a bacterial infection caused by Corynebacterium pseudotuberculosis (C. pseudotuberculosis) that affects sheep and goats, leading to abscess formation in their lymph nodes. The present study aimed to isolate and identify C. pseudotuberculosis from CLA in smallholder sheep and goats, and determine the resistance patterns, virulence, and resistance genes of the isolates. Additionally, genotypic and phylogenetic analysis of the isolates was conducted using ERIC-PCR and DNA sequencing techniques. A cross-sectional study examined 220 animals (130 sheep and 90 goats) from 39 smallholder flocks for clinical signs of CLA. Fifty-four (24.54%) animals showed CLA-compatible lesions, confirmed by C. pseudotuberculosis isolation and PCR identification. Sheep had a lower infection rate of CLA (18.46%) compared with goats (33.3%). Antimicrobial susceptibility testing of 54 C. pseudotuberculosis isolates to 24 antimicrobial drugs revealed that they were 100% resistant to bacitracin and florfenicol, while none of the isolates were resistant to norfloxacin. A high resistance rate was observed for penicillin and erythromycin (92.6% each). Interestingly, 16.7% of C. pseudotuberculosis isolates recovered from sheep showed vancomycin resistance. Molecular characterization of C. pseudotuberculosis isolates revealed that PLD, PIP, and FagA virulence genes were present in all examined isolates. However, the FagB, FagC, and FagD genes were detected in 24 (100%), 20 (83%), and 18 (75%) of the sheep isolates, and 26 (87%), 26 (87%), and 18 (60%) of the goat isolates, respectively. The ß-lactam resistance gene was present in all isolates. Furthermore, 83% of the sheep isolates carried the aminoglycoside (aph(3″)-lb), chloramphenicol (cat1), and bacitracin (bcrA) resistance genes. Among the isolates recovered from goats, 73% were found to contain macrolides (ermX), sulfonamide (sul1), and bacitracin (bcrA) resistance genes. It is worrisome that the glycopeptide (vanA) resistance gene was detected in 8% of the sheep isolates as a first report. ERIC-PCR genotyping of 10 multi-drug-resistant C. pseudotuberculosis isolates showed a high similarity index of 83.6% between isolates from sheep and goats. Nucleotide sequence analysis of partial 16S rRNA sequences of C. pseudotuberculosis revealed 98.83% similarity with biovar Ovis of globally available reference sequences on the Genbank database. Overall, our findings might indicate that C. pseudotuberculosis infection in smallholders in Egypt might be underestimated despite the significant financial impact on animal husbandry and potential health hazards it poses. Moreover, this study highlights the importance of implementing a sustainable control strategy and increasing knowledge and awareness among smallholder breeders to mitigate the economic impact of CLA.

Seroprevalence and risk factors of tropical theileriosis in smallholder asymptomatic large ruminants in Egypt.

Knowledge of the prevalence and epidemiological determinants of tropical theileriosis in large ruminants, particularly in the asymptomatic carrier, is crucial for designing and implementing effective host-specific control measures. This study aimed to estimate the seroprevalence of tropical theileriosis in asymptomatic cattle and water buffaloes and identify the potential risk factors of theileriosis in large ruminants raised under smallholder-production system in Egypt. A cross-sectional study was conducted in five districts of the Sharkia governorate from March 2019 to February 2020. In total, 350 serum samples were collected from cattle and water buffaloes under smallholder-production system and tested for Theileria annulata antibodies using the indirect antibody fluorescence test (IFAT). Data on species, host characteristics, presence of ticks, season, and districts were collected at sampling using a questionnaire. A multivariable mixed-effects logistic regression model was built to determine the potential risk factors associated with T. annulate seropositivity of the animals. The overall apparent seroprevalence of T. annulata in 350 tested animals was 70%. In the univariable analyses, cattle compared to buffaloes, younger animals compared to older ones, animals with ticks on their bodies, and warmer seasons were all associated with a higher likelihood of seropositive results in the study population while sex of the animals was not associated with seropositivity. The final multivariable model showed that animals with ticks on their bodies had 3.5× higher odds of seropositivity than those with no ticks (P < 0.001), and warmer seasons were associated with the higher odds of infection compared to winter (P = 0.003). The high seroprevalence of tropical theileriosis in the study region indicates that the disease is endemic among smallholders of large ruminants. The identified risk factors of T. annulata-seropositivity in asymptomatic carrier animals provides evidence-based guidance for adopting effective intervention measures.

Genetic characterization of upper respiratory tract virome from nonvaccinated Egyptian cow-calf operations.

Bovine respiratory disease (BRD) is the costliest complex disease affecting the cattle industry worldwide, with significant economic losses. BRD pathogenesis involves several interactions between microorganisms, such as bacteria and viruses, and management factors. The present study aimed to characterize the nasal virome from 43 pooled nasal swab samples collected from Egyptian nonvaccinated cow-calf operations with acute BRD from January to February 2020 using metagenomic sequencing. Bovine herpesvirus-1 (BHV-1), first detection of bovine herpesvirus-5 (BHV-5), and first detection of bovine parvovirus-3 (BPV-3) were the most commonly identified in Egyptian cattle. Moreover, phylogenetic analysis of glycoprotein B revealed that the BHV-1 isolate is closely related to the Cooper reference strain (genotype 1.1), whereas the BHV-5 isolate is closely related to the reference virus GenBank NP_954920.1. In addition, the whole-genome sequence of BPV-3 showed 93.02% nucleotide identity with the reference virus GenBank AF406967.1. In this study, several DNA viruses, such as BHV-1 and first detection BHV-5, and BPV-3, were detected and may have an association with the BRD in Egyptian cattle. Therefore, further research, including investigating more samples from different locations to determine the prevalence of detected viruses and their contributions to BRD in cattle in Egypt, is needed.

Virulence Determinants and Plasmid-Mediated Colistin Resistance mcr Genes in Gram-Negative Bacteria Isolated From Bovine Milk.

A major increase of bacterial resistance to colistin, a last-resort treatment for severe infections, was observed globally. Using colistin in livestock rearing is believed to be the ground of mobilized colistin resistance (mcr) gene circulation and is of crucial concern to public health. This study aimed to determine the frequency and virulence characteristics of colistin-resistant Gram-negative bacteria from the milk of mastitic cows and raw unpasteurized milk in Egypt. One hundred and seventeen strains belonging to Enterobacteriaceae (n = 90), Pseudomonas aeruginosa (n = 10), and Aeromonas hydrophila (n = 17) were screened for colistin resistance by antimicrobial susceptibility testing. The genetic characteristics of colistin-resistant strains were investigated for mcr-1-9 genes, phylogenetic groups, and virulence genes. Moreover, we evaluated four commonly used biocides in dairy farms for teat disinfection toward colistin-resistant strains. Multidrug-resistant (MDR) and extensive drug-resistant (XDR) phenotypes were detected in 82.91% (97/117) and 3.42% (4/117) of the isolates, respectively. Of the 117 tested isolates, 61 (52.14%) were colistin resistant (MIC >2 mg/L), distributed as 24/70 (34.29%) from clinical mastitis, 10/11 (90.91%) from subclinical mastitis, and 27/36 (75%) from raw milk. Of these 61 colistin-resistant isolates, 47 (19 from clinical mastitis, 8 from subclinical mastitis, and 20 from raw milk) harbored plasmid-borne mcr genes. The mcr-1 gene was identified in 31.91%, mcr-2 in 29.79%, mcr-3 in 34.04%, and each of mcr-4 and mcr-7 in 2.13% of the colistin-resistant isolates. Among these isolates, 42.55% (20/47) were E. coli, 21.28% (10/47) A. hydrophila, 19.12% (9/47) K. pneumoniae, and 17.02% (8/47) P. aeruginosa. This is the first report of mcr-3 and mcr-7 in P. aeruginosa. Conjugation experiments using the broth-mating technique showed successful transfer of colistin resistance to E. coli J53-recipient strain. Different combinations of virulence genes were observed among colistin-resistant isolates with almost all isolates harboring genes. Hydrogen peroxide has the best efficiency against all bacterial isolates even at a low concentration (10%). In conclusion, the dissemination of mobile colistin resistance mcr gene and its variants between MDR- and XDR-virulent Gram-negative isolates from dairy cattle confirms the spread of mcr genes at all levels; animals, humans, and environmental, and heralds the penetration of the last-resort antimicrobial against MDR bacteria. Consequently, a decision to ban colistin in food animals is urgently required to fight XDR and MDR bacteria.

Whole-Genome Sequencing of Gram-Negative Bacteria Isolated From Bovine Mastitis and Raw Milk: The First Emergence of Colistin mcr-10 and Fosfomycin fosA5 Resistance Genes in Klebsiella pneumoniae in Middle East.

Antimicrobial resistance is a major concern in the dairy industry. This study investigated the prevalence, antimicrobial resistance phenotypes, and genome sequencing of Gram-negative bacteria isolated from clinical (n = 350) and subclinical (n = 95) bovine mastitis, and raw unpasteurized milk (n = 125). Klebsiella pneumoniae, Aeromonas hydrophila, Enterobacter cloacae (100% each), Escherichia coli (87.78%), and Proteus mirabilis (69.7%) were the most prevalent multidrug-resistant (MDR) species. Extensive drug-resistance (XDR) phenotype was found in P. mirabilis (30.30%) and E. coli (3.33%) isolates. Ten isolates (four E. coli, three Klebsiella species and three P. mirabilis) that displayed the highest multiple antibiotic resistance (MAR) indices (0.54-0.83), were exposed to whole-genome sequencing (WGS). Two multilocus sequence types (MLST): ST2165 and ST7624 were identified among the sequenced E. coli isolates. Three E. coli isolates (two from clinical mastitis and one from raw milk) belonging to ST2165 showed similar profile of plasmid replicon types: IncFIA, IncFIB, IncFII, and IncQ1 with an exception to an isolate that contained IncR, whereas E. coli ST7624 showed a different plasmid profile including IncHI2, IncHI2A, IncI1α, and IncFII replicon types. ResFinder findings revealed the presence of plasmid-mediated colistin mcr-10 and fosfomycin fosA5 resistance genes in a K. pneumoniae (K1) isolate from bovine milk. Sequence analysis of the reconstructed mcr-10 plasmid from WGS of K1 isolate, showed that mcr-10 gene was bracketed by xerC and insertion sequence IS26 on an IncFIB plasmid. Phylogenetic analysis revealed that K1 isolate existed in a clade including mcr-10-harboring isolates from human and environment with different STs and countries [United Kingdom (ST788), Australia (ST323), Malawi (ST2144), Myanmar (ST705), and Laos (ST2355)]. This study reports the first emergence of K. pneumoniae co-harboring mcr-10 and fosA5 genes from bovine milk in the Middle East, which constitutes a public health threat and heralds the penetration of the last-resort antibiotics. Hence, prudent use of antibiotics in both humans and animals and antimicrobial surveillance plans are urgently required.

Environmental Streptococcus uberis Associated with Clinical Mastitis in Dairy Cows: Virulence Traits, Antimicrobial and Biocide Resistance, and Epidemiological Typing.

Mastitis remains a serious problem for dairy animals. The misappropriation of antimicrobial agents helps accelerate resistance, which poses a serious challenge in controlling environmental S. uberis infection. Here, we study the virulence attributes, antimicrobial and biocide resistance, and epidemiological typing of S. uberis recovered from bovine clinical mastitis in dairy farms of diverse hygienic interventions in Egypt. The overall S. uberis infection rate was 20.59%; all were multidrug-resistant (MDR). The sua gene was the most frequent virulence gene (42.02%), followed by pauA (40.57%), cfu (21.73%), skc (20.28%), and opp (11.59%). The erm(B) gene served as the predominant antimicrobial-resistant gene (75.36%), followed by fexA (52.63%) and tet(M), blaZ, and aac(6')aph(2″) genes (46.38% each). Of note, 79.71%, 78.26%, and 18.84% of S. uberis isolates harbored qacED1, qacC/D, and qacA/B genes, respectively. All analyzed isolates were S. uberis type I by their unique RFLP-PCR pattern. In conclusion, the sustained presence of pauA and sua genes throughout the investigated farms contributes to a better understanding of the bacterium's pathogenicity. Furthermore, MDR coupled with the existence of biocide resistance genes indicates the importance of S. uberis surveillance and the prudent use of antimicrobials in veterinary clinical medicine to avoid the dissemination of antimicrobial resistance.

Characterization of Foot and Mouth Disease Virus Serotype SAT-2 in Swamp Water Buffaloes (Bubalus bubalis) under the Egyptian Smallholder Production System.

Spontaneous mutations are a common characteristic of the foot and mouth disease virus (FMDV), leading to wide antigenic variations resulting in the emergence of new topotypes and lineages of FMDV, which contributes to occasional vaccination failures. The objectives of the present study were to genetically characterize FMDV isolated from water buffaloes and study the biochemical and histopathological indicators of infected animals. Fifty-four water buffaloes of both sexes and different ages suffered from acute symptoms of FMD were clinically examined and randomly selected for inclusion in this study. Oral desquamated epithelial and oropharyngeal fluid samples have been tested for FMDV by reverse transcriptase PCR (RT-PCR). Tissue and serum samples were also collected from the diseased buffaloes and subjected to histopathological and biochemical analysis. Our findings showed that all examined samples were confirmed to be positive to FMDV serotype SAT-2 and were adjusted to be responsible for the recent disease outbreak in this study. Phylogenetic analysis revealed that the circulating viruses were of the SAT-2 serotype, closely related to the lineage of lib12, topotype VII, with 98.9% identity. The new lineage of SAT-2 showed a high virulence resulting in the deaths of water buffaloes due to heart failure, confirmed by high serum levels of inflammatory and cardiac markers, including haptoglobin, ceruloplasmin, cardiac troponin I and creatine phosphokinase-MB, indicating an unfavorable FMD-infection prognosis. In conclusion, we document the presence of new incursions circulating in water buffalo populations in Egypt in early 2019, explaining the high morbidity rate of FMD outbreak in early 2019. Furthermore, the newly identified serotype SAT-2 lib12 lineage, topotype VII, showed an aggressive pattern in water buffaloes of the smallholder production system.

Sensitivity and specificity of piroplasm indirect fluorescent antibody test and PCR for Theileria annulata infection in clinically asymptomatic large ruminants using Bayesian latent class analysis.

There is limited information about the accuracy of molecular and serological diagnostic assays for tropical theileriosis in asymptomatic carrier large ruminants. This study has estimated the sensitivity (Se) and specificity (Sp) of PCR and an indirect fluorescent antibody test (IFAT) in the diagnosis of tropical theileriosis in cattle and buffaloes via a Bayesian latent class analysis (BLCA) framework. Blood samples were collected from 70 cattle and water buffaloes (Bubalus bubalis) raised under a smallholder production system in different Egyptian localities. T. annulata infection status was detected by PCR, and IFAT and the test results were subjected to BLCA without assuming the existence of a reference test. Our findings showed that the performance of PCR was superior to that of IFAT. PCR showed a higher Se [0.83 (95% PCI: 0.63-0.98)] in comparison to IFAT [0.72 (95% PCI: 0.68-0.75)]. Similarly, PCR showed a higher Sp [0.95 (95% PCI: 0.77-1.00)] than IFAT [0.82 (95% PCI: 0.80-0.84)]. Se and Sp of the two tests did not differ by species implying that the diagnostics' performance for T. annulata infection in bovines is the same regardless of the species under consideration. In conclusion, PCR outperforms IFAT in the detection of T. annulata infection and can thus be applied to routine control of tropical theileriosis in endemic situations where cattle and buffaloes are kept under traditional smallholder production systems.

Antimicrobial and antibiofilm potentials of cinnamon oil and silver nanoparticles against Streptococcus agalactiae isolated from bovine mastitis: new avenues for countering resistance.

BACKGROUND: Streptococcus agalactiae (S. agalactiae) is a contagious pathogen of bovine mastitis. It has financial implications for the dairy cattle industry in certain areas of the world. Since antimicrobial resistance increases in dairy farms, natural antimicrobials from herbal origins and nanoparticles have been given more attention as an alternative therapy. Hence, this study reported the antimicrobial and antibiofilm potentials of cinnamon oil, silver nanoparticles (AgNPs), and their combination against multidrug-resistant (MDR) S. agalactiae recovered from clinical bovine mastitis in Egypt. RESULTS: Our findings revealed that 73% (146/200) of the examined milk samples collected from dairy cows with clinical mastitis were infected with Streptococci species. Of these, 9.59% (14/146) were identified as S. agalactiae and categorized as MDR. S. agalactiae isolates expressed four virulence genes (Hyl, cylE, scpB, and lmb) and demonstrated an ability to produce biofilms. Cinnamon oil showed high antimicrobial (MICs ≤0.063 µg /mL) and antibiofilm (MBIC50 = 4 µg/mL) potentials against planktonic and biofilms of S. agalactiae isolates, respectively. However, AgNPs showed reasonable antimicrobial (MICs ≤16 µg/mL) and relatively low antibiofilm (MBIC50 = 64 µg/mL) activities against screened isolates. Synergistic antimicrobial or additive antibiofilm interactions of cinnamon oil combined with AgNPs were reported for the first time. Scanning electron microscope (SEM) analysis revealed that biofilms of S. agalactiae isolates treated with cinnamon oil were more seriously damaged than observed in AgNPs cinnamon oil combination. Moreover, reverse transcriptase quantitative polymerase chain reaction (RT-qPCR) showed that cinnamon oil exerted a remarkable down-regulation of pili biosynthesis genes (pilA and pilB) and their regulator (rogB) against S. agalactiae biofilms, meanwhile the AgNPs cinnamon oil combination demonstrated a lower efficacy. CONCLUSIONS: This is an in vitro preliminary approach that documented the antibiofilm potential of cinnamon oil and the inhibitory activity of cinnamon oil and its combination with AgNPs against MDR S. agalactiae recovered from clinical mastitis. Further in vivo studies should be carried out in animal models to provide evidence of concept for implementing these alternative candidates in the treatment of dairy farms infected by streptococcal mastitis in the future.

Molecular characterization of a novel Camelus dromedarius papillomavirus.

Papillomaviruses affect both human and non-human hosts. In camels, papillomatosis is caused by Camelus dromedarius papillomavirus type 1 and 2 (CdPV1 and CdPV2, respectively). In late 2018, an outbreak of camelpox occurred in a herd of fattening camels in Egypt. Several animals were found to be co-infected with camelpox and camel papillomaviruses. The morbidity with papillomatosis was 35 %. The infection was confirmed by PCR then Illumina sequencing revealed the presence of a complete genome of two CdPVs. One of these was CdPV1 (MT130101) and the other was a putative novel virus, tentatively named as CdPV3 (MT130100). Seven ORFs and a long upstream regulatory region were identified in the genomes of both viruses. Pairwise comparisons of L1 gene revealed 98.92 % nt identity between MT130101/CdPV1/Egypt/2018 and HQ912790/CdPV1/Sudan/2009 with 100 % coverage. However, MT130100/CdPV3/ Egypt/2018 showed only 68.99 % nt identity with the closest genome HQ912791/CdPV2/Sudan/2009. Phylogenetic analyses indicated that CdPV1 and CdPV3 belonged to the genus Deltapapillomavirus. These results should be useful for future CdPVs molecular surveillance and construction of evolutionary characteristics of this virus.

Epidemiological and ultrasonographic investigation of bovine fascioliasis in smallholder production system in Eastern Nile Delta of Egypt.

Regular updating of our knowledge on the epidemiological determinants of bovine fascioliasis is necessary to increase the awareness of the disease's significance and subsequently, improve the control measures. The objectives of this study were (1) to estimate the prevalence of bovine fascioliasis, and identify the association of epidemiological characteristics under traditional householders' production systems, (2) to describe the association between the clinical picture, Fasciola spp. egg count and hepatobiliary ultrasonography findings. In total, 270 faecal samples were examined microscopically for the presence or absence of Fasciola spp. egg, using the sedimentation-flotation method. Copro-positive animals were subjected to ultrasonographic examination. Overall prevalence of copro-positive animals was 27.4% (22.4-33.0%, 95% CI). The final multivariate analysis showed that there was a significant association between fascioliasis and animal species (P < 0.03), and administration of anthelmintic (P < 0.0001). Cattle have a less chance of being positive to Fasciola spp. by 0.55 (95% CI: 0.30 - 0.99) compared to water buffaloes. Administration of anthelmintic to animals on a regular basis decreased the risk of copro-positivity to Fasciola spp by 0.17 (95% CI: 0.07 - 0.36) compared to animals received anthelmintic on an irregular basis. Infected animals having different Fasciola spp. egg burden revealed different clinical symptoms associated with hepatobiliary changes on ultrasonographic examination ranged from normal hepatic parenchyma and bile system in low faecal egg load to hyperechogenic hepatic parenchyma, hyperechogenic with distal shadowing bile duct, and distended gallbladder in high faecal egg load of Fasciola spp. In conclusion, the prevalence of bovine fascioliasis is high under the traditional household's production system. Regular administration of anthelmintic significantly reduces the animal's chance of being copro-positive to Fasciola spp. Ultrasound poses a valuable prognostic technique for assessment of bovine fascioliasis.

Sporadic cases of lumpy skin disease among cattle in Sharkia province, Egypt: Genetic characterization of lumpy skin disease virus isolates and pathological findings.

BACKGROUND AND AIM: Lumpy skin disease (LSD) is a highly infectious viral disease upsetting cattle, caused by LSD virus (LSDV) within the family Poxviridae. Sporadic cases of LSD have been observed in cattle previously vaccinated with the Romanian sheep poxvirus (SPPV) vaccine during the summer of 2016 in Sharkia province, Egypt. The present study was undertaken to perform molecular characterization of LSDV strains which circulated in this period as well as investigate their phylogenetic relatedness with published reference capripoxvirus genome sequences. MATERIALS AND METHODS: A total of 82 skin nodules, as well as 5 lymph nodes, were collected from suspect LSD cases, and the virus was isolated in embryonated chicken eggs (ECEs). LSD was confirmed by polymerase chain reactions amplification of the partial and full-length sequences of the attachment and G-protein-coupled chemokine receptor (GPCR) genes, respectively, as well as a histopathological examination of the lesions. Molecular characterization of the LSDV isolates was conducted by sequencing the GPCR gene. RESULTS: Characteristic skin nodules that covered the whole intact skin, as well as lymphadenopathy, were significant clinical signs in all suspected cases. LSDV isolation in ECEs revealed the characteristic focal white pock lesions dispersed on the chorioallantoic membranes. Histopathologic examination showed characteristic eosinophilic intracytoplasmic inclusion bodies within inflammatory cell infiltration. Phylogenetic analysis revealed that the LSDV isolates were clustered together with other African and European LSDV strains. In addition, the LSDV isolates have a unique signature of LSDVs (A11, T12, T34, S99, and P199). CONCLUSION: LSDV infections have been detected in cattle previously vaccinated with Romanian SPPV vaccine during the summer of 2016 and making the evaluation of vaccine efficacy under field conditions necessary.

Diagnostic performance of molecular and conventional methods for identification of dermatophyte species from clinically infected Arabian horses in Egypt.

BACKGROUND: Rapid and accurate identification of dermatophytes is crucial for the effective control of disease outbreaks. Current methods based on culture and microscopic characteristics may require weeks before positive identification is made. OBJECTIVES: To (i) identify the most common pathogenic dermatophytes affecting Arabian horses; (ii) compare the performance of direct microscopy (DM), culture, PCR using hair samples (PCRhair ) and PCR based on culture isolates (PCRculture ) for the diagnosis of dermatophytosis. METHODS: Samples of hair and crusts of skin lesions from Arabian horses were collected on a monthly basis by scraping skin of affected horses. Samples were divided into three portions: the first portion was used for microscopic examination, the second for culture and the third portion for PCR amplification of intergenic spacer (ITS) regions. RESULTS: Out of 200 horses examined, 70 (35%) showed cutaneous lesions characteristic of dermatophytosis. DM revealed that 70.4% were positive for fungal elements and 85.7% were culture positive. The identified species were Microsporum canis, Trichophyton verrucosum, T. mentagrophytes var. mentagrophytes and M. equinum. Among 25 selected samples, 64, 92, 91.3 and 52% were positive for dermatophytes, as determined by DM, culture, PCRculture and PCRhair , respectively. CONCLUSIONS: The dermatophytes M. canis, T. verrucosum, T. mentagrophytes var. mentagrophytes and M. equinum were the most common cause of dermatophytosis in Arabian horses. Although the number of samples was small, the ITS-based PCR may be a useful diagnostic tool when combined with culture.