Effects of Eimeria acervulina infection on the luminal and mucosal microbiota of the cecum and ileum in broiler chickens.

Coccidiosis, an intestinal disease caused by Eimeria parasites, is responsible for major losses in the poultry industry by impacting chicken health. The gut microbiota is associated with health factors, such as nutrient exchange and immune system modulation, requiring understanding on the effects of Eimeria infection on the gut microbiota. This study aimed to determine the effects of Eimeria acervulina infection on the luminal and mucosal microbiota of the cecum (CeL and CeM) and ileum (IlL and IlM) at multiple time points (days 3, 5, 7, 10, and 14) post-infection. E. acervulina infection decreased evenness in CeL microbiota at day 10, increased richness in CeM microbiota at day 3 before decreasing richness at day 14, and decreased richness in IlL microbiota from day 3 to 10. CeL, CeM, and IlL microbiota differed between infected and control birds based on beta diversity at varying time points. Infection reduced relative abundance of bacterial taxa and some predicted metabolic pathways known for short-chain fatty acid production in CeL, CeM, and IlL microbiota, but further understanding of metabolic function is required. Despite E. acervulina primarily targeting the duodenum, our findings demonstrate the infection can impact bacterial diversity and abundance in the cecal and ileal microbiota.

Integrative and Conjugative Elements and Prophage DNA as Carriers of Resistance Genes in Erysipelothrix rhusiopathiae Strains from Domestic Geese in Poland.

Goose erysipelas is a serious problem in waterfowl breeding in Poland. However, knowledge of the characteristics of Erysipelothrix rhusiopathiae strains causing this disease is limited. In this study, the antimicrobial susceptibility and serotypes of four E. rhusiopathiae strains from domestic geese were determined, and their whole-genome sequences (WGSs) were analyzed to detect resistance genes, integrative and conjugative elements (ICEs), and prophage DNA. Sequence type and the presence of resistance genes and transposons were compared with 363 publicly available E. rhusiopathiae strains, as well as 13 strains of other Erysipelothrix species. Four strains tested represented serotypes 2 and 5 and the MLST groups ST 4, 32, 242, and 243. Their assembled circular genomes ranged from 1.8 to 1.9 kb with a GC content of 36-37%; a small plasmid was detected in strain 1023. Strains 1023 and 267 were multidrug-resistant. The resistance genes detected in the genome of strain 1023 were erm47, tetM, and lsaE-lnuB-ant(6)-Ia-spw cluster, while strain 267 contained the tetM and ermB genes. Mutations in the gyrA gene were detected in both strains. The tetM gene was embedded in a Tn916-like transposon, which in strain 1023, together with the other resistance genes, was located on a large integrative and conjugative-like element of 130 kb designated as ICEEr1023. A minor integrative element of 74 kb was identified in strain 1012 (ICEEr1012). This work contributes to knowledge about the characteristics of E. rhusiopathiae bacteria and, for the first time, reveals the occurrence of erm47 and ermB resistance genes in strains of this species. Phage infection appears to be responsible for the introduction of the ermB gene into the genome of strain 267, while ICEs most likely play a key role in the spread of the other resistance genes identified in E. rhusiopathiae.

In Vitro Evaluation of Phytobiotic Mixture Antibacterial Potential against Enterococcus spp. Strains Isolated from Broiler Chicken.

Enterococcus spp. are normal intestinal tract microflorae found in poultry. However, the last decades have shown that several species, e.g., Enterococcus cecorum, have become emerging pathogens in broilers and may cause numerous losses in flocks. In this study, two combinations (H1 and H2) of menthol, 1,8-cineol, linalool, methyl salicylate, γ-terpinene, p-cymene, trans-anethole, terpinen-4-ol and thymol were used in an in vitro model, analyzing its effectiveness against the strains E. cecorum, E. faecalis, E. faecium, E. hirae and E. gallinarum isolated from broiler chickens from industrial farms. To identify the isolated strains classical microbiological methods and VITEK 2 GP cards were used. Moreover for E. cecorum a PCR test was used.. Antibiotic sensitivity (MIC) tests were performed for all the strains. For the composition H1, the effective dilution for E. cecorum and E. hirae strains was 1:512, and for E. faecalis, E. faecium and E. gallinarum, 1:1024. The second mixture (H2) showed very similar results with an effectiveness at 1:512 for E. cecorum and E. hirae and 1:1024 for E. faecalis, E. faecium and E. gallinarum. The presented results suggest that the proposed composition is effective against selected strains of Enterococcus in an in vitro model, and its effect is comparable to classical antibiotics used to treat this pathogen in poultry. This may suggest that this product may also be effective in vivo and provide effective support in the management of enterococcosis in broiler chickens.

Dust sprinkling as an effective method for infecting layer chickens with wild-type Salmonella Typhimurium and changes in host gut microbiota.

Role of dust in Salmonella transmission on chicken farms is not well characterised. Salmonella Typhimurium (ST) infection of commercial layer chickens was investigated using a novel sprinkling method of chicken dust spiked with ST and the uptake compared to a conventional oral infection. While both inoculation methods resulted in colonisation of the intestines, the Salmonella load in liver samples was significantly higher at 7 dpi after exposing chicks to sprinkled dust compared to the oral infection group. Infection of chickens using the sprinkling method at a range of doses showed a threshold for colonisation of the gut and organs as low as 1000 CFU/g of dust. Caecal content microbiota analysis post-challenge showed that the profiles of chickens infected by the sprinkling and oral routes were not significantly different; however, both challenges induced differences when compared to the uninfected negative controls. Overall, the study showed that dust sprinkling was an effective way to experimentally colonise chickens with Salmonella and alter the gut microbiota than oral gavage at levels as low as 1000 CFU/g dust. This infection model mimics the field scenario of Salmonella infection in poultry sheds. The model can be used for future challenge studies for effective Salmonella control.

Prevalence and antimicrobial susceptibility of Salmonella enteritidis and Salmonella typhimurium isolated from hen eggs and quail eggs in Karaj, Iran.

BACKGROUND AND AIM: Different Salmonella serotypes are considered one of the most important food pathogens in the world. Poultry meat and eggs are the primary carriers of Salmonella in human populations. This study aimed to estimate the Salmonella enteritidis and Salmonella typhimurium contamination rates of retail hen and quail eggs in Karaj, Iran. Moreover, the antimicrobial resistance patterns of the strains were evaluated, and the efficiency of the standard culture method and multiplex polymerase chain reaction (m-PCR) were compared. MATERIALS AND METHODS: In this descriptive cross-sectional study over 1 year (Jan-Dec 2022), 150 commercial and 150 backyard hen eggs and 300 commercial quail eggs, without cracks and fractures, were collected randomly from best selling groceries in Karaj city. All samples were examined for Salmonella contamination independently by standard culture and m-PCR approaches. A standard disc diffusion method was employed to assess the antimicrobial susceptibility of the strains against 18 antimicrobial agents. RESULTS: Out of 300 examined eggs, 2 S. enteritidis strains were isolated from the shell of backyard hen eggs. The same serotype was also detected in the contents of one of these two eggs. One S. typhimurium was isolated from the shell of a commercial hen egg. Overall, the Salmonella contamination of the shell and contents was 1% and 0.3%, respectively. Salmonella was not isolated from the eggshells or the contents of the quail eggs. There was complete agreement between the results of m-PCR and the standard culture methods. Among the 18 tested antibiotics, the highest resistance was recorded for colistin (100%), followed by nalidixic acid (75%). CONCLUSION: As most Salmonella spp. are associated with human food poisoning, continuous surveillance is required to effectively reduce the risk posed by contaminated poultry eggs. Furthermore, mandatory monitoring of antimicrobial use on Iranian poultry farms is recommended.

Prevalence, geographic distribution and risk factors of Eimeria species on commercial broiler farms in Guangdong, China.

BACKGROUND: Coccidiosis is one of the most frequently reported diseases in chickens, causing a significant economic impact on the poultry industry. However, there have been no previous studies evaluating the prevalence of this disease in broiler farms in Guangdong province. Therefore, this study aims to conduct an epidemiological investigation into the occurrence of Eimeria species and associated risk factors in intensive management conditions across four regions in Guangdong province, China. A total of 394 fecal samples were collected from 89 broiler farms in Guangdong province. The prevalence of Eimeria species infection was determined using PCR, and the occurrence of Clostridium perfringens type A was assessed using quantitative real-time PCR. RESULTS: The results showed an overall prevalence of 98.88% (88/89) at the farm level and 87.06% (343/394) at the flock level. All seven Eimeria species were identified, with E. acervulina (72.53%; 64/89), E. tenella (68.54%; 61/89), and E. mitis (66.29%; 59/89) at the farm level, and E. acervulina (36.55%; 144/394), E. mitis (35.28%; 139/394), and E. tenella (34.01%; 134/394) at the flock level. The predominant species combination observed was a co-infection of all seven Eimeria species (6.74%; 6/89), followed by a combination of E. acervulina, E. tenella, E. mitis, E. necatrix, E. brunetti, and E. maxima (5.62%, 5/89). A combination of E. acervulina, E. tenella, E. mitis, E. necatrix, E. brunetti, and E. praecox (4.49%; 4/89) was also observed at the farm level. Furthermore, the study identified several potential risk factors associated with the prevalence of Eimeria species, including farm location, chicken age, drinking water source, control strategy, and the presence of C. perfringens type A were identified as potential risk factors associated with prevalence of Eimeria species. Univariate and multivariate analyses revealed a significant association between E. necatrix infection and both grower chickens (OR = 10.86; 95% CI: 1.92-61.36; p < 0.05) and adult chickens (OR = 24.97; 95% CI: 4.29-145.15; p < 0.001) compared to starter chickens at the farm level. Additionally, farms that used groundwater (OR = 0.27; 95% CI: 0.08-0.94; p < 0.05) were less likely to have E. maxima compared to those that used running water. At the flock level, the prevalence of E. tenella was significantly higher in the Pearl River Delta (OR = 2.48; 95% CI: 1.0-6.15; p = 0.05) compared to eastern Guangdong. Interestingly, flocks with indigenous birds were less likely to have E. brunetti (OR = 0.48; 95% CI: 0.26-0.89; p < 0.05) compared to flocks with indigenous crossbred birds. Furthermore, flocks that used anticoccidial drugs (OR = 0.09; 95% CI: 0.03-0.31; p < 0.001) or a combination of vaccines and anticoccidial drugs (OR = 0.06; 95% CI: 0.01-0.25; p < 0.001) were less likely to be positive for E. tenella compared to flocks that only used vaccines. Finally, flocks with C. perfringens type A infection were significantly more likely to have E. necatrix (OR = 3.26; 95% CI: 1.96-5.43; p < 0.001), E. tenella (OR = 2.14; 95% CI: 1.36-3.36; p < 0.001), E. brunetti (OR = 2.48; 95% CI: 1.45-4.23; p < 0.001), and E. acervulina (OR = 2.62; 95% CI: 1.69-4.06; p < 0.001) compared to flocks without C. perfringens type A. CONCLUSIONS: This study conducted an investigation on the prevalence, distribution, and risk factors associated with Eimeria species infection in broiler chickens in Guangdong. The farm-level prevalence of Eimeria species was higher than the previous prevalence figures for other areas and countries. E. brunetti was identified at higher prevalence in Guangdong than previously survived prevalence in different regions in China. Farm location, chicken age, drinking water source, control strategy, and the presence of C. perfringens type A were considered as potential risk factors associated with prevalence of Eimeria species. It is imperative to underscore the necessity for further surveys to delve deeper into the occurrence of Eimeria species under intensive management conditions for different flock purposes.

Evaluation of the safety and efficacy of the novel Mycoplasma gallisepticum vaccine, Vaxsafe MG304, after spray-vaccination of 1-day-old specific pathogen-free chicks.

Mycoplasma gallisepticum causes chronic respiratory disease in poultry. A novel vaccine, Vaxsafe MG304 (the ts-304 strain), has greater protective efficacy in chickens than the Vaxsafe MG (strain ts-11) vaccine when delivered by eye drop at 3 weeks of age. Applying this vaccine in the hatchery to 1-day-old birds, using mass administration methods, would improve animal welfare and reduce labour costs associated with handling individual birds. This study assessed the protection provided by vaccination with Vaxsafe MG304 after administration to 1-day-old chicks. Chicks were administered a single dose of the vaccine to assess the efficacy of either a high dose (107.0 colour changing units, CCU) or a low dose (105.7 CCU) after eye drop or spray (in water or gel) administration against experimental challenge with virulent M. gallisepticum strain Ap3AS at 7 weeks of age. The vaccine was able to colonise the palatine cleft of chicks after vaccination by eye drop (at both doses) or by spray (in water or gel) (at the high dose). The high dose of vaccine, when delivered by eye drop or spray, was shown to be safe and induced a serological response and protective immunity (as measured by tracheal mucosal thickness and air sac lesion scores) against challenge. Vaccination of 1-day-old chicks with Vaxsafe MG304 by eye drop induced protective immunity equivalent to vaccination at 3 weeks of age. Vaxsafe MG304 was also protective when applied by both coarse- and gel spray methods at the higher dose and is therefore a suitable live attenuated vaccine for use in 1-day-old chicks.

The impact of different levels of functional oil supplementation in combination with salinomycin on growth performance and intestinal microbiota of broilers undergoing Eimeria challenge: An analysis of dynamics.

The effect of salinomycin sodium alone and in combination with functional oils on performance and microbiota of broiler infected Eimeria were evaluated. 512 broilers were randomly assigned to 4 treatments (8 replicates, 16 birds/pen): a Control group (any additives); Ionophore group: salinomycin supplementation at 66 ppm (SS66); Ionophore +0.075% Functional oil (FO) group (SS66 + FO supplementation at 750 ppm); and Ionophore +0.10% FO group (SS66 + FO supplementation at 1000 ppm). At 14 days of age, birds were gavaged with 1 mL of a saline solution containing sporulated oocysts of E. tenella, E. acervulina and E. maxima. Performance indices were measured weekly. At 28 days, intestinal content was collected for microbiota analysis. Broilers of Control group presented the worst performance indices. Broilers of Ionophore + FO (0.075% and 0.10%) groups exhibited a higher BW at 28 days of age. The supplementation of Ionophore +0.075% FO resulted in a higher relative proportion of Firmicutes and a lower proportion of Actinobacteria in the ileum-jejunum. Lactobacillaceae was the dominant family in the jejunal, and ileal microbiotas of broilers fed diets supplemented with Ionophore, Ionophore +0.075% FO and Ionophore +0.10% FO. The supplementation of ionophore yielded higher numbers of Lactobacillaceae, Enterobactereaceae and Cloritridiaceae in the cecal. Ionophore associated with FO controlled the Lactobacillaceae, Enterobactereaceae and Cloritridiaceae families present in the cecum. Therefore, the combination of salinomycin with functional oil showed synergistic effect on performance and modulation of intestinal microbiota of broilers challenged with Eimeria.

Survival of highly related ESBL- and pAmpC- producing Escherichia coli in broiler farms identified before and after cleaning and disinfection using cgMLST.

BACKGROUND: Broiler chickens are frequently colonized with Extended-Spectrum Beta-Lactamase- (ESBL-) and plasmid mediated AmpC Beta-Lactamase- (pAmpC-) producing Enterobacterales, and we are confronted with the potential spread of these resistant bacteria in the food chain, in the environment, and to humans. Research focused on identifying of transmission routes and investigating potential intervention measures against ESBL- and pAmpC- producing bacteria in the broiler production chain. However, few data are available on the effects of cleaning and disinfection (C&D) procedures in broiler stables on ESBL- and pAmpC- producing bacteria. RESULTS: We systematically investigated five broiler stables before and after C&D and identified potential ESBL- and pAmpC- colonization sites after C&D in the broiler stables, including the anteroom and the nearby surrounding environment of the broiler stables. Phenotypically resistant E. coli isolates grown on MacConkey agar with cefotaxime were further analyzed for their beta-lactam resistance genes and phylogenetic groups, as well as the relation of isolates from the investigated stables before and after C&D by whole genome sequencing. Survival of ESBL- and pAmpC- producing E. coli is highly likely at sites where C&D was not performed or where insufficient cleaning was performed prior to disinfection. For the first time, we showed highly related ESBL-/pAmpC- producing E. coli isolates detected before and after C&D in four of five broiler stables examined with cgMLST. Survival of resistant isolates in investigated broiler stables as well as transmission of resistant isolates from broiler stables to the anteroom and surrounding environment and between broiler farms was shown. In addition, enterococci (frequently utilized to detect fecal contamination and for C&D control) can be used as an indicator bacterium for the detection of ESBL-/pAmpC- E. coli after C&D. CONCLUSION: We conclude that C&D can reduce ESBL-/pAmpC- producing E. coli in conventional broiler stables, but complete ESBL- and pAmpC- elimination does not seem to be possible in practice as several factors influence the C&D outcome (e.g. broiler stable condition, ESBL-/pAmpC- status prior to C&D, C&D procedures used, and biosecurity measures on the farm). A multifactorial approach, combining various hygiene- and management measures, is needed to reduce ESBL-/pAmpC- E. coli in broiler farms.

Antimicrobial effects of laurel extract, laurel essential oil, zahter extract, and zahter essential oil on chicken wings contaminated with Salmonella Typhimurium.

BACKGROUND: This study aimed to evaluate the antimicrobial effects of zahter extract, zahter essential oil, laurel extract, and laurel essential oil on Salmonella Typhimurium inoculated on chicken wings. METHODS: A total of 10 groups, including eight study groups and two control groups were formed, consisting of zahter extract and zahter essential oil and laurel extract and laurel essential oil in different proportions. In the study, laurel extract at 6.4% and 12.8% concentrations, laurel essential oil at 0.2% and 0.4% concentrations, zahter extract at 0.2% and 0.4% concentrations, and zahter essential oil at 0.2% and 0.4% concentrations were used. RESULTS: The broth microdilution method was used to evaluate the antimicrobial activity of the extract and essential oils on the S. Typhimurium. Minimum inhibitory concentrations of the extracts and essential oils used in the study against S. Typhimurium were determined. The highest inhibitory effect on S. Typhimurium was observed in the 0.4% laurel essential oil group. It was determined that the inhibitory effect increased as the concentration of laurel essential oil increased. In addition, the antimicrobial activity of zahter essential oil is less inhibitory than the laurel extract, laurel essential oil, and zahter extract. CONCLUSION: According to the results of this study, it has been revealed that extracts and essential oils obtained from zahter and laurel plants, which have been shown to be natural antimicrobial, can be used in foods as an alternative to chemical additives. To develop research results, the applicability of these extracts and essential oils in different foodstuffs should be examined using different ingredients and concentrations.

Efficacy evaluation of hydrogen peroxide disinfectant based zinc oxide nanoparticles against diarrhea causing Escherichia coli in ruminant animals and broiler chickens.

Different strains of Escherichia coli that exhibit genetic characteristics linked to diarrhea pose a major threat to both human and animal health. The purpose of this study was to determine the prevalence of pathogenic Escherichia coli (E. coli), the genetic linkages and routes of transmission between E. coli isolates from different animal species. The efficiency of disinfectants such as hydrogen peroxide (H2O2), Virkon®S, TH4+, nano zinc oxide (ZnO NPs), and H2O2-based zinc oxide nanoparticles (H2O2/ZnO NPs) against isolated strains of E. coli was evaluated. Using 100 fecal samples from different diarrheal species (cow n = 30, sheep n = 40, and broiler chicken n = 30) for E. coli isolation and identification using the entero-bacterial repetitive intergenic consensus (ERIC-PCR) fingerprinting technique. The E. coli properties isolated from several diarrheal species were examined for their pathogenicity in vitro. Scanning electron microscopy (SEM), high-resolution transmission electron microscopy (HR-TEM), Fourier-transform infrared spectrum (FT-IR), X-ray diffraction (XRD), zeta potential, and particle size distribution were used for the synthesis and characterization of ZnO NPs and H2O2/ZnO NPs. The broth macro-dilution method was used to assess the effectiveness of disinfectants and disinfectant-based nanoparticles against E. coli strains. Regarding the results, the hemolytic activity and Congo red binding assays of pathogenic E. coli isolates were 55.3 and 44.7%, respectively. Eleven virulent E. coli isolates were typed into five ERIC-types (A1, A2, B1, B2, and B3) using the ERIC-PCR method. These types clustered into two main clusters (A and B) with 75% similarity. In conclusion, there was 90% similarity between the sheep samples' ERIC types A1 and A2. On the other hand, 89% of the ERIC types B1, B2, and B3 of cows and poultry samples were comparable. The H2O2/ZnO NPs composite exhibits potential antibacterial action against E. coli isolates at 0.04 mg/ml after 120 min of exposure.

Effect of combined infection with Salmonella and influenza virus on their respective proliferation in chicken embryonated eggs.

Background: Salmonella is a major food-borne bacterial pathogen that causes food poisoning related to the consumption of eggs, milk, and meat. Food safety in relation to Salmonella is particularly important for eggs because their shells as well as their contents can be a source of contamination. Chicken can also be infected with influenza virus, but it remains unclear how co-infection of Salmonella and influenza virus affect each other. Aim: The potential influence of co-infection of Salmonella and influenza virus was examined. Methods: Salmonella Abony and influenza virus were injected into chicken embryonated eggs. After incubation, proliferation of Salmonella and influenza virus was measured using a direct culture assay for bacteria and an enzyme-linked immunosorbent assay for influenza virus, respectively. Results: Our findings indicate that the number of colony-forming units (CFUs) of Salmonella did not vary between chicken embryonated eggs co-infected with influenza A virus and Salmonella-only infected eggs. Furthermore, we found the proliferation of influenza A or B virus was not significantly influenced by co-infection of the eggs with Salmonella. Conclusion: These results suggest that combined infection of Salmonella with influenza virus does not affect each other, at least in terms of their proliferation.

Antimicrobial resistance patterns and genes of Campylobacter jejuni isolated from chickens in Pasuruan, Indonesia.

Background: Poultry is one of the most prominent sources of Campylobacter jejuni, which is also a major means of transmission to people. Campylobacter jejuni contamination in chicken meat comes from chicken feces because it naturally exists in the intestines of chickens. Aim: The purpose of this study is to identify the antibiotic resistance patterns and genes of C. jejuni, which was found in chickens in Pasuruan, Indonesia. Methods: The samples used in this study were 200 contents of the small intestine of broiler chickens from 40 farms in Pasuruan Regency. The enriched sample was streaked on the selective media of modified charcoal cefoperazone deoxycholate agar containing the CCDA selective supplement. Antimicrobial susceptibility test utilizing the Kirby-Bauer diffusion test method in accordance with Clinical and Laboratory Standards Institute standards. The polymerase chain reaction (PCR) method was used to detect the (hipO), which encodes the C. jejuni strain, fluoroquinolone resistance (gyrA), beta-lactam resistance (blaOXA-61), and tetracycline resistance (tetO) genes. Results: The findings revealed a 14% (28/200) prevalence of C. jejuni in the small intestine of broiler chickens. These isolates showed high resistance to enrofloxacin (92.9%). All isolates (100%) were susceptible to amoxicillin-clavulanate. The PCR results showed all C. jejuni isolates (100%) detected the gyrA gene, 96.4% detected the blaOXA-61 gene, and 50% detected the tetO gene. Conclusion: The findings of antimicrobial resistance at a high level from the small intestine of broiler chickens illustrate the potential threat to human health. To lessen the effects now and in the future, coordinated and suitable action is needed, as well as steps to guarantee the poultry industry's economic survival and public health insurance.

Combined application of bacteriophages with a competitive exclusion culture and carvacrol with organic acids can reduce Campylobacter in primary broiler production.

For reducing Campylobacter (C.) in the food production chain and thus the risk to the consumer, the combined application of different measures as a multiple-hurdle approach is currently under discussion. This is the first study to investigate possible synergistic activities in vivo, aiming at reducing intestinal C. jejuni counts by administering (i) bacteriophages (phages) in combination with a competitive exclusion (CE) product and (ii) carvacrol combined with organic acids. The combined application of the two selected phages (Fletchervirus phage NCTC 12673 and Firehammervirus phage vB_CcM-LmqsCPL1/1) and the CE product significantly reduced C. jejuni loads by 1.0 log10 in cecal and colonic contents as well as in cloacal swabs at the end of the trial (33 and 34 days post hatch). The proportion of bacterial isolates showing reduced phage susceptibility ranged from 10.9% (isolates from cecal content) to 47.8% (isolates from cloacal swabs 32 days post hatch) for the Fletchervirus phage, while all tested isolates remained susceptible to the Firehammervirus phage. The use of carvacrol combined with an organic acid blend (sorbic acid, benzoic acid, propionic acid, and acetic acid) significantly reduced Campylobacter counts by 1.0 log10 in cloacal swabs on day 30 only.

Occurrence, antibiotic resistance profiles and associated risk factors of Klebsiella pneumoniae in poultry farms in selected districts of Somalia Reginal State, Ethiopia.

BACKGROUND: Klebsiella pneumoniae is an opportunistic infection that causes production losses and death in the chicken industry. A cross-sectional study was conducted on exotic chicken breeds reared at the Jigjiga poultry farm from November 2022 to May 2023 to estimate the occurrence, associated risk factors, and antimicrobial susceptibility profiles of Klebsiella pneumoniae. The chickens were selected using systematic random sampling techniques. A total of 384 cloacal swabs were collected aseptically and transported to the laboratory for analysis. For statistical analysis, STATA® version 14.0 statistical software was used. RESULTS: From 384 examined faecal samples, 258 (67.2%) prevalences of Klebsiella pneumoniae were found. Furthermore, the association of the study's risk factors with the prevalence of Klebsiella pneumoniae was explored, and no statistically significant association was identified between sex and age. Nonetheless, relative prevalence at the age level was higher in chickens aged 12 months (67.6%) and Sasso breeds (90.0%). Similarly, male chickens and those raised for meat and egg production had a high prevalence rate of 72.5 and 75.8%, respectively. A total of 30 isolated Klebsiella pneumoniae colonies were tested in vitro for antibiotic sensitivity for six drugs, and it was shown that Klebsiella pneumoniae is moderately sensitive to Penicillin G (43.3%) while having higher resistance to Oxytetracycline (80.0%). CONCLUSIONS: The current findings revealed that the research area had the highest prevalence of Klebsiella pneumoniae, and the isolates were resistant to commonly used drugs in the study area. Thus, a long-term intervention plan, thorough research to determine a nationwide status, as well as further multi-drug resistance patterns and molecular characterization, were urged.

A comprehensive study of colisepticaemia progression in layer chickens applying novel tools elucidates pathogenesis and transmission of Escherichia coli into eggs.

Colisepticaemia caused by avian pathogenic Escherichia coli (APEC) is a challenging disease due to its high economic importance in poultry, dubious pathogenesis and potential link with zoonosis and food safety. The existing in vitro studies can't define hallmark traits of APEC isolates, suggesting a paradigm shift towards host response to understand pathogenesis. This study investigated the comprehensive pathological and microbial progression of colisepticaemia, and transmission of E. coli into eggs using novel tools. In total 48 hens were allocated into three groups and were inoculated intratracheally with ilux2-E. coli PA14/17480/5-/ovary (bioluminescent strain), E. coli PA14/17480/5-/ovary or phosphate buffered saline. Infection with both strains led to typical clinical signs and lesions of colibacillosis as in field outbreaks. Based on lung histopathology, colisepticaemia progression was divided into four disease stages as: stage I (1-3 days post infection (dpi)), stage II (6 dpi), stage III (9 dpi) and stage IV (16 dpi) that were histologically characterized by predominance of heterophils, mixed cells, pyogranuloma, and convalescence, respectively. As disease progressed, bacterial colonization in host organs also decreased, revealed by the quantification of bacterial bioluminescence, bacteriology, and quantitative immunohistochemistry. Furthermore, immunofluorescence, immunohistochemistry, and bacteria re-isolation showed that E. coli colonized the reproductive tract of infected hens and reached to egg yolk and albumen. In conclusion, the study provides novel insights into the pathogenesis of colisepticemia by characterizing microbial and pathological changes at different disease stages, and of the bacteria transmission to table eggs, which have serious consequences on poultry health and food safety.

Pathovars, occurrence, and characterization of plasmid-mediated quinolone resistance in diarrheal Escherichia coli isolated from farmers and farmed chickens in Tunisia and Nigeria.

The poultry industry is a very important agricultural and industrial sector in Tunisia and Nigeria, with little information about occurrence of diarrheagenic Escherichia coli in the farmers and chickens. This study aimed to detect the prevalence of diarrheal E. coli in humans and poultry and to investigate plasmid-mediated quinolone resistance (PMQR) genes in both countries. Seventy-four isolates of E. coli were studied; nine different virulence genes were screened by PCR. Serotyping was performed only for pathotypes as well as the determining of antibiotic resistance profiles against 21 antibiotics. PMQR genes were investigated by PCR. EAEC was the most abundant pathotype (37/74; 50%) in human and chicken isolates, whereas single EHEC and EPEC (1/74, 1.35%) pathotypes were detected in Tunisia and Nigeria, respectively. About 17 (45.95%) quinolones/fluoroquinolones-resistant isolates were detected, from which the following PMQR genes were detected: aac(6')-Ib-cr (8/17, 47.05%), qepA (6/17, 35.29%), qnrA + qnrB (2/17, 11.76%), and qnrS gene (1/17, 5.88%). Our findings highlight high occurrence of EAEC pathotype in Tunisia and Nigeria, more frequent than EPEC and EHEC. Additionally, all E. coli pathotypes isolated from different sources (humans, poultry) showed resistance to several antibiotics, which are in use as therapeutic choices in Tunisia and Nigeria.

Evolution and genomic profile of Salmonella enterica serovar Gallinarum biovar Pullorum isolates from Brazil.

Salmonella enterica subspecies enterica serovar Gallinarum biovar Pullorum (S. Pullorum) is a pathogenic bacterium that causes Pullorum disease (PD). PD is an acute systemic disease that affects young chickens, causing white diarrhea and high mortality. Although many sanitary programs have been carried out to eradicate S. Pullorum, PD outbreaks have been reported in different types of birds (layers, broilers, breeders) worldwide. This study aimed to evaluate the evolution and genetic characteristics of S. Pullorum isolated from PD in Brazil. Phylogenetic analysis of S. Pullorum genomes sequenced in this study and available genomic databases demonstrated that all isolates from Brazil are from sequence type 92 (ST92) and cluster into two lineages (III and IV). ColpVC, IncFIC(FII), and IncFII(S) were plasmid replicons frequently found in the Brazilian lineages. Two resistance genes (aac(6')-Iaa, conferring resistance to aminoglycoside, disinfecting agents, and antiseptics (mdf(A)) and tetracycline (mdf(A)) were detected frequently. Altogether, these results are important to understand the circulation of S. Pullorum and, consequently, to develop strategies to reduce losses due to PD.

Evolución y perfil genómico de aislados de Salmonella enterica serovar Gallinarum biovar Pullorum de Brasil. Salmonella enterica subespecie enterica serovar Gallinarum biovar Pullorum (S. Pullorum) es una bacteria patógena que causa la enfermedad de Pullorum (EP). La EP es una enfermedad sistémica aguda que afecta a los pollos jóvenes causando diarrea blanca y alta mortalidad. Aunque se han llevado a cabo muchos programas sanitarios para erradicar S. Pullorum, se han informado brotes de EP en diferentes tipos de aves (ponedoras, pollos de engorde, reproductoras) en todo el mundo. Este estudio tuvo como objetivo evaluar la evolución y las características genéticas de S. Pullorum aislado de EP en Brasil. El análisis filogenético de los genomas de S. Pullorum secuenciados en este estudio y las bases de datos genómicas disponibles demostraron que todos los aislamientos de Brasil son del tipo de secuencia 92 (ST92) y se agrupan en dos linajes (III y IV). ColpVC, IncFIC (FII) e IncFII(S) fueron replicones de plásmidos frecuentemente encontrados en los linajes brasileños. Dos genes de resistencia (aac(6')-Iaa, que confiere resistencia a aminoglucósidos, desinfectantes y antisépticos (mdf(A)), y tetraciclina (mdf(A)) fueron detectados con frecuencia. En conjunto, estos resultados son importantes para comprender la circulación de S. Pullorum y, en consecuencia, desarrollar estrategias para reducir las pérdidas por EP.

In silico and PCR Screening for a Live Attenuated Salmonella Typhimurium Vaccine Strain.

The application of live attenuated Salmonella Typhimurium vaccines has significantly helped control Salmonella in poultry products. Because the U.S. Department of Agriculture-Food Safety Inspection Service (USDA-FSIS) scores all Salmonella as positive, regardless of serovar, attenuated vaccine strains that are identified at processing contribute negatively toward Salmonella performance standards. This study was designed to determine the incidence of a live attenuated Salmonella serovar Typhimurium vaccine identified in broiler products by FSIS and to develop a PCR assay for screening of isolates. Salmonella Typhimurium short-read sequences from broiler samples uploaded to the National Center for Biotechnology Information (NCBI) Pathogen Detection database by the USDA-FSIS from 2016 to 2022 were downloaded and assembled. These were analyzed using the Basic Local Alignment Search Tool (BLAST) with a sequence unique to field strains, followed by a sequence unique to the vaccine strain. The PCR assays were developed against field and vaccine strains by targeting transposition events in the crp and cya genes and validated by screening Salmonella serovar Typhimurium isolates. Between 2016 and 2022, 1708 Salmonella Typhimurium isolates of chicken origin were found in the NCBI Pathogen Detection database, corresponding to 7.99% of all Salmonella identified. Of these, 104 (5.97%) were identified as the vaccine strain. The PCR assay differentiated field strains from the vaccine strain when applied to isolates and was also able to detect the vaccine strain from DNA isolated from mixed serovar overnight Salmonella enrichment cultures. Live attenuated Salmonella vaccines are a critical preharvest tool for Salmonella control and are widely used in industry. With forthcoming regulations that will likely focus on Salmonella Typhimurium, along with other serovars, there is a need to distinguish between isolates belonging to the vaccine strain and those that are responsible for causing human illness.

Detección in silico y por PCR de una cepa vacunal viva atenuada de Salmonella Typhimurium. La aplicación de vacunas vivas atenuadas contra Salmonella Typhimurium ha ayudado significativamente a controlar Salmonella en productos avícolas. Debido a que el Servicio de Inspección de Seguridad Alimentaria del Departamento de Agricultura de los Estados Unidos. (USDA-FSIS) califica todas las Salmonella como positivas, independientemente del serovar. Las cepas atenuadas de la vacuna que se identifican en el procesamiento contribuyen negativamente a los estándares de desempeño de Salmonella. Este estudio fue diseñado para determinar la incidencia de una vacuna viva atenuada de Salmonella serovar Typhimurium identificada en productos de pollo de engorde por el FSIS y para desarrollar un ensayo de PCR para la detección de aislados. Se recolectaron y ensamblaron secuencias de lectura corta de Salmonella Typhimurium de muestras de pollos de engorde introducidas en la plataforma de detección de patógenos del Centro Nacional de Información Biotecnológica (NCBI) por el USDA-FSIS entre los años 2016 al 2022. Estos se analizaron utilizando la herramienta de búsqueda de alineación local básica con una secuencia exclusiva para las cepas de campo, seguida de una secuencia exclusiva para la cepa vacunal. Los ensayos de PCR se desarrollaron contra cepas de campo y vacunales centrándose en eventos de transposición en los genes crp y cya y se validaron mediante la detección de aislados de Salmonella serovar Typhimurium. Entre 2016 y 2022, se encontraron 1708 aislados de Salmonella Typhimurium de origen avícola en el sistema de detección de patógenos del NCBI, lo que corresponde al 7.99 % de todas las Salmonellas identificadas. De ellas, 104 (5.97%) fueron identificadas como cepa vacunal. El ensayo de PCR diferenció las cepas de campo de la cepa de la vacuna cuando se aplicó a los aislados y también fue capaz de detectar la cepa de la vacuna a partir del ADN aislado de cultivos de enriquecimiento por toda la noche de Salmonella con serovares mixtos. Las vacunas vivas atenuadas contra Salmonella son una herramienta fundamental para el control de Salmonella y se utilizan ampliamente en la industria. Con las próximas regulaciones que probablemente se centrarán en Salmonella Typhimurium, junto con otros serovares, es necesario distinguir entre los aislados que pertenecen a la cepa vacunal y los que son responsables de causar enfermedades humanas.

Immunization of turkeys with Clostridium septicum alpha toxin-based recombinant subunit proteins can confer protection against experimental Clostridial dermatitis.

Clostridial dermatitis (CD), caused by Clostridium septicum, is an emerging disease of increasing economic importance in turkeys. Currently, there are no effective vaccines for CD control. Here, two non-toxic domains of C. septicum alpha toxin, namely ntATX-D1 and ntATX-D2, were identified, cloned, and expressed in Escherichia coli as recombinant subunit proteins to investigate their use as potential vaccine candidates. Experimental groups consisted of a Negative control (NCx) that did not receive C. septicum challenge, while the adjuvant-only Positive control (PCx), ntATX-D1 immunization (D1) and ntATX-D2 immunization (D2) groups received C. septicum challenge. Turkeys were immunized subcutaneously with 100 µg of protein at 7, 8 and 9 weeks of age along with an oil-in-water nano-emulsion adjuvant, followed by C. septicum challenge at 11 weeks of age. Results showed that while 46.2% of birds in the PCx group died post-challenge, the rate of mortality in D1- or D2-immunization groups was 13.3%. The gross and histopathological lesions in the skin, muscle and spleen showed that the disease severity was highest in PCx group, while the D2-immunized birds had significantly lower lesion scores when compared to PCx. Gene expression analysis revealed that PCx birds had significantly higher expression of pro-inflammatory cytokine genes in the skin, muscle and spleen than the NCx group, while the D2 group had significantly lower expression of these genes compared to PCx. Peripheral blood cellular analysis showed increased frequencies of activated CD4+ and/or CD8+ cells in the D1 and D2-immunized groups. Additionally, the immunized turkeys developed antigen-specific serum IgY antibodies. Collectively, these findings indicate that ntATX proteins, specifically the ntATX-D2 can be a promising vaccine candidate for protecting turkeys against CD and that the protection mechanisms may include downregulation of C. septicum-induced inflammation and increased CD4+ and CD8+ cellular activation.