First seroprevalence and molecular identification report of Brucella canis among dogs in Greater Cairo region and Damietta Governorate of Egypt.

Background and Aim: Given the rise in stray and imported dogs in Egypt over the past 5 years, it is surprising that no report of Brucella canis infection in dogs or humans has been documented in Egypt's published papers. This study aimed to detect the presence of antibodies against the rough (B. canis) and smooth Brucellae among dogs in Egypt and to characterize the Brucella species circulating in dogs. Materials and Methods: Blood samples (n = 449) were collected from owned and stray dogs in the Greater Cairo region (n = 309) and Damietta governorate (n = 140). The apparent, true, and total seroprevalence of canine brucellosis caused by B. canis infection were calculated using the 2-mercaptoethanol tube agglutination test (2-ME TAT) and rapid slide agglutination test (RSAT). We used the rose Bengal test (RBT) and the buffered acidified plate antigen test (BAPAT) to check the serum samples from dogs for the presence of antibodies against smooth Brucellae. Three polymerase chain reaction (PCR) assays - Bruce-ladder PCR, B. canis species-specific PCR (BcSS-PCR), and Abortus Melitensis Ovis Suis (AMOS)-PCR - were used to determine the Brucella species in the buffy coats of the serologically positive dogs. Results: The overall apparent and true prevalence of B. canis infection in dogs were estimated to be 3.8% and 13.2%. The estimated true prevalence in stray dogs (15%) was higher than in owned dogs (12.5%). The BAPAT and the RBT using smooth antigens revealed that 11 (2.4%) and 9 (2%) were positive. Bruce-ladder PCR targeting eryC, ABC, and Polysaccharide deacetylase genes was able to identify B. canis in nine out of 17 buffy coat samples. AMOS-PCR identified the eight undetermined Brucella species by Bruce-ladder PCR as Brucella abortus (n = 4) and Brucella melitensis (n = 4). To exclude the presence of Brucella suis, a one-step species-specific BcSS-PCR was performed and specifically amplified all B. canis DNA (n = 9) the same as did the Bruce-ladder PCR. Conclusion: To the best of our knowledge, this is the first report of B. canis detection in dogs in Egypt. Molecular identification of B. abortus and B. melitensis in the Egyptian canines highlights the role of stray dogs in brucellosis remerging in Brucellosis-free dairy farms. Brucella canis infection can be diagnosed specifically with the one-step BcSS-PCR. The obtained results set-an-alarm to the veterinary authorities to launch plans to control this disease in dogs.

Enteral administration of a simulated amniotic fluid in preventing feeding intolerance in very low birthweight neonates: A randomized controlled trial.

OBJECTIVES: Feeding intolerance (FI) is a common finding in preterm neonates. Enteral administration of different forms of amniotic fluid (AF) has been tried for treating FI in high-risk neonates. Simulated amniotic fluid (SAF) is a solution with a similar electrolyte composition to human AF. The aim of this study was to examine whether enteral administration of SAF would improve feeding tolerance in very low birthweight (VLBW) neonates. METHODS: Forty VLBW neonates were randomized to either SAF or placebo (total daily dose 20 mL/kg/d-1 divided every 3 h) to their milk for a maximum of 7 d. Neonates with major congenital anomalies, those in whom early feeding was contraindicated, and those treated with parental erythropoietin and/or human granulocyte stimulating factor were excluded. The primary outcome was the total amount of enteral feeds reached by day 7. Secondary outcomes were incidence of FI and necrotizing enterocolitis (NEC). Study intervention was stopped on completing 7 d, reaching enteral feeds of 100 mL/kg/d-1, or the appearance of any sign of FI or NEC. RESULTS: All neonates tolerated the test solution well. The SAF group reached significantly larger volume and higher calories on days 3 and 7 (P < 0.05 for all). No statistical differences were seen between the two groups in incidence of FI (P = 0.311), NEC (P = 0.429), mortality (P = 0.632), length of stay (P = 0.744), or weight gain on day 10 (P = 0.389). Baseline hematologic parameters showed no statistical differences before or after enteral administration (P > 0.05). CONCLUSION: Results of the present study demonstrated that SAF solution might improve feeding tolerance in VLBW babies without evidence of its systemic absorption. Larger multicenter randomized studies are recommended.

Whole-Genome Sequencing for Tracing the Genetic Diversity of Brucella abortus and Brucella melitensis Isolated from Livestock in Egypt.

Brucellosis is a highly contagious zoonosis that occurs worldwide. Whole-genome sequencing (WGS) has become a widely accepted molecular typing method for outbreak tracing and genomic epidemiology of brucellosis. Twenty-nine Brucella spp. (eight B. abortus biovar 1 and 21 B. melitensis biovar 3) were isolated from lymph nodes, milk, and fetal abomasal contents of infected cattle, buffaloes, sheep, and goats originating from nine districts in Egypt. The isolates were identified by microbiological methods and matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS). Differentiation and genotyping were confirmed using multiplex PCR. Illumina MiSeq® was used to sequence the 29 Brucella isolates. Using MLST typing, ST11 and ST1 were identified among B. melitensis and B. abortus, respectively. Brucella abortus and B. melitensis isolates were divided into two main clusters (clusters 1 and 2) containing two and nine distinct genotypes by core-genome SNP analysis, respectively. The genotypes were irregularly distributed over time and space in the study area. Both Egyptian B. abortus and B. melitensis isolates proved to be genomically unique upon comparison with publicly available sequencing from strains of neighboring Mediterranean, African, and Asian countries. The antimicrobial resistance mechanism caused by mutations in rpoB, gyrA, and gyrB genes associated with rifampicin and ciprofloxacin resistance were identified. To the best of our knowledge, this is the first study investigating the epidemiology of Brucella isolates from livestock belonging to different localities in Egypt based on whole genome analysis.

Risk factors and Molecular genotyping of Brucella melitensis strains recovered from humans and their owned cattle in Upper Egypt.

Brucellosis is a zoonosis that has a devastating impact on the economy and public health, particularly in the Middle East, including Egypt. This study aimed to define risk factors associated with brucellosis in humans and in their cattle in Fayoum governorate - Upper Egypt. Also, molecular genotyping of recovered Brucella isolates from human cases and their cattle to assess the potential cross-species transmission in the study region. Data were obtained via double matched case-control studies for brucellosis in humans (106 cases and 160 controls) and in their cattle (78 cattle cases and 105 cattle controls). The results of multivariate regression analysis revealed that predictors of human brucellosis were animal-related occupations (OR 2.1, P 0.02), previous infection in other household members (OR 3.2, P 0.007), eating home-made soft cheese (OR 2.3, P 0.03), and exposure to cattle abortions (OR 6.9, P < 0.001). For cattle, predictors of brucellosis were maturity ≥2 years of age (OR 2.9, P 0.01), ≥2 animals reared by the same household (OR 3.7-6.9, P ≤ 0.001), and recent abortion (OR 15.2, P 0.01). Twelve Brucella isolates were recovered from eight human cases (7.5%, 8/106) and four cattle cases (6.2%, 4/65). All isolates were B. melitensis biovar 3. Analysis of the IS711 gene sequence revealed complete homology (100%) between isolates. Six virulence genes were utilized for virutyping: virB (100%), omp25 (100%), amiC (100%), ure (91.7%), wbkA (91.7%), and bvfA (75%). Virutyping revealed four virutypes: V1 (lack bvfA, 16.7%), V2 (harbored all genes, 66.7%), V3 (lack wbkA, 8.3%), and V4 (lack wbkA and ure, 8.3%). Repetitive extragenic palindromic PCR (REP-PCR) typing revealed two REP types. Combined REP-PCR/virulence genotyping revealed five different genotypes (G1-G5) for the detected isolates and a unique genotype for the reference strain (G6, B. melitensis bv3 Ether). Human and cattle isolates from the same household had matched genotypes. In conclusion, there were widespread risk factors among the cases studied. Health education for high-risk groups is essential for disease prevention, and combined REP-PCR/virulence genotyping is a quick tool for traceability, particularly in developing countries endemic with brucellosis as Egypt.

Validation of real-time polymerase chain reaction versus conventional polymerase chain reaction for diagnosis of brucellosis in cattle sera.

BACKGROUND AND AIM: Different polymerase chain reaction (PCR) techniques have and are still being used for the direct detection of Brucella DNA in serum samples of different animal species and humans without being validated or properly validated, resulting in discrepancies. Thus, this study aimed to evaluate the diagnostic performance of the TaqMan Real-Time-PCR (RT-PCR) targeting the bcsp31 gene versus conventional PCR for the accurate diagnosis of brucellosis at the genus level in cattle sera. MATERIALS AND METHODS: One hundred and eighty-four serum samples were collected from bacteriologically positive and negative cows with ages ranging from 1 to 5 years old at some infected private farms in the Nile Delta under quarantine measures as well as brucellosis free farms. These samples were classified into four groups after serological diagnosis and investigated by TaqMan RT-PCR and conventional PCR targeting the IS711 gene for Brucella DNA detection. The diagnostic performance characteristics of both PCR techniques were estimated considering the bacteriological results as a gold standard. RESULTS: TaqMan RT-PCR revealed superiority over conventional PCR; it was able to detect Brucella DNA in 95% (67/70) and 89% (25/28) of the cattle sera samples belonging to Group 1 (serologically and bacteriologically positive) and Group 2 (serologically negative but bacteriologically positive), respectively. On evaluating the diagnostic performance, TaqMan RT-PCR showed superior diagnostic sensitivity (93.9%), diagnostic specificity (88.4%), performance index (182.3), almost perfect kappa agreement (0.825±0.042), strong positive correlation (r=0.826), high accuracy based on the receiver operating characteristic (ROC) curve, and area under the ROC curve (0.911) at p<0.05 and CI of 95%. CONCLUSION: A cattle serum sample is not the metric of choice for targeting Brucella genomic DNA by conventional PCR. The time-saving and rapid TaqMan RT-PCR method revealed a better diagnostic performance in the detection of Brucella DNA in cattle sera. Such performance offered by TaqMan RT-PCR may be considered a step toward the possibility of using such technology in the direct differentiation between Brucella-infected and -vaccinated cattle immunized by smooth vaccines from cattle sera using primers specific for such vaccines.

Gut priming with bovine colostrum and T regulatory cells in preterm neonates: a randomized controlled trial.

BACKGROUND: Necrotizing enterocolitis (NEC) and neonatal sepsis are still considered major problems, especially in formula-fed preterm neonates. This study aimed to investigate the effect of bovine colostrum on T regulatory cells, NEC, and late-onset sepsis in preterm neonates ≤34 weeks. METHODS: This prospective double-blind randomized controlled trial was conducted on 80 preterm infants who were randomly assigned to either the bovine colostrum group (n = 32) or control group (n = 48). T lymphocytes and their subsets, necrotizing enterocolitis, late-onset sepsis (LOS) and its severity, feeding tolerance, growth, length of hospital stay, and mortality were documented. RESULTS: The bovine colostrum group showed higher follow-up levels of CD4+CD25+ FOXP3+ T lymphocyte % (FOXP3 Tregs). FOXP3 Tregs and its difference in change levels between baseline and follow-up were considered as the most related factors to the bovine colostrum. Bovine colostrum group showed positive trends for reduction of sepsis severity and mortality with no significant difference in the incidence of NEC, LOS, and length of hospital stay. CONCLUSIONS: Preterm neonates who received bovine colostrum showed a higher FOXP3 Treg level. IMPACT: Bovine colostrum has no significant effect on the incidence of necrotizing enterocolitis. FOXP3 T regulatory cells and their increased level between baseline and follow-up is considered as the most influencing factors related to the bovine colostrum. Positive trends were noted for reduction of sepsis severity and concomitant mortality, but the study lacked the power to assess these outcomes.

High serum APRIL levels strongly correlate with disease severity in pediatric atopic eczema.

BACKGROUND: Expression of the tumor necrosis factor (TNF) family member APRIL (a proliferation-inducing ligand) is low in normal tissues but is elevated in various autoimmune diseases. OBJECTIVES: This study explored serum APRIL in children with atopic eczema (AE) during flare and quiescence. METHODS: A case-control study including 50 patients with AE and 40 control subjects was conducted. The severity of AE was assessed according to each of the Leicester Sign Score (LSS), the Simple Scoring System (SSS), the SCORing of Atopic Dermatitis (SCORAD) index, and the Objective SCORAD. Serum measurements of APRIL, total immunoglobulin E, and lactate dehydrogenase were obtained in all subjects. Data were obtained during both flare and quiescence in AE subjects. Data were analyzed using the Mann-Whitney U-test and Pearson's correlation coefficient. P-values of <0.05 were considered to indicate statistical significance. RESULTS: Serum APRIL levels were significantly elevated in children with AE in comparison with control subjects during both flare and quiescence (P < 0.0001 for both). Serum APRIL levels during AE flare and quiescence were positively correlated (P < 0.001). Serum APRIL levels and scores on each of the LSS, SSS, and SCORAD showed significant positive correlations (P < 0.01 for all). CONCLUSIONS: Serum APRIL levels were significantly elevated in children with AE during both flare and quiescence. This confirms the importance of APRIL in the pathophysiology of pediatric AE. Serum APRIL is a reliable marker of the severity of AE in children. APRIL may be a new target in the treatment of AE.

Gentamicin extended interval regimen and ototoxicity in neonates.

OBJECTIVES: To assess the extended interval regimen gentamicin associated ototoxicity in neonatal intensive care unit using hearing tests. METHODS: Two hundred and twenty neonates admitted to neonatal intensive care were assessed; 110 neonates who had received gentamicin and 110 neonates who had not received gentamicin served as control group. Gentamicin group were further subdivided according to the duration of treatment into 50 neonates who had received gentamicin for 5 days or less and 60 neonates who had received gentamicin for more than 5 days. TEOAEs were used for hearing screening. Auditory brain response was performed 3 months later for failed cases to confirm the hearing impairment. RESULTS: Three neonates failed TEOAEs screening in each group but hearing impairment was confirmed in one neonate only (0.9%) in each group (gentamicin and control groups). Neonates who received gentamicin for more than 5 days showed comparable results as regard TEOAEs or ABR results with those who received gentamicin for 5 days or less, and control group. CONCLUSIONS: Extended interval dosing of gentamicin therapy in neonates does not increase the incidence of hearing loss. This suggests that hearing loss in neonatal intensive care unit may be attributed to factors other than gentamicin treatment.

Effect of caffeine on preterm infants' cerebral cortical activity: an observational study.

OBJECTIVE: Our first aim was to investigate the effects of caffeine on preterm infants' respiratory functions and brain cortical activity (conventional and amplitude-integrated electroencephalography (cEEG and aEEG)). Secondary aim was to study its long-term effects on respiratory system and electroencephalographic maturation by 36 weeks post-menstrual age. METHODS: Prospective observational study on 33 consecutively admitted preterm infants less than 34-weeks-gestation. cEEG and aEEG, cardiopulmonary and sleep state were recorded in 20 preterm infants, before, during and 2-hours after intravenous (IV) caffeine (caffeine Group), and for 13 preterms (control group). Both groups were subjected to assessment of cerebral cortical maturation by cEEG and aEEG at 36-weeks post-menstrual age as an outcome measure. RESULTS: IV caffeine administration significantly increased heart rate (p = 0.000), mean arterial blood pressure (p = 0.000), capillary oxygen saturation (p = 0.003), arousability (p = 0.000) and aEEG continuity (p = 0.002) after half an hour. No clinical seizures were recorded and non-significant difference was found in electrographic seizures activity in cEEG. At 36-weeks post-conceptional age, NICU stay was significantly longer in controls (p = 0.022). aEEG score was significantly higher in caffeine group than the control group, (p = 0.000). CONCLUSIONS: Caffeine increases preterm infants' cerebral cortical activity during infusion and results in cerebral cortical maturation at 36weeks, without increase in seizure activity.