Necrotic enteritis (NE), mainly induced by the pathogens of Clostridium perfringens and coccidia, causes huge economic losses with limited intervention options in the poultry industry. This study investigated the role of specific bile acids on NE development. Day-old broiler chicks were assigned to six groups: noninfected, NE, and NE with four bile diets of 0.32% chicken bile, 0.15% commercial ox bile, 0.15% lithocholic acid (LCA), or 0.15% deoxycholic acid (DCA). The birds were infected with Eimeria maxima at day 18 and C. perfringens at day 23 and 24. The infected birds developed clinical NE signs. The NE birds suffered severe ileitis with villus blunting, crypt hyperplasia, epithelial line disintegration, and massive immune cell infiltration, while DCA and LCA prevented the ileitis histopathology. NE induced severe body weight gain (BWG) loss, while only DCA prevented NE-induced BWG loss. Notably, DCA reduced the NE-induced inflammatory response and the colonization and invasion of C. perfringens compared to NE birds. Consistently, NE reduced the total bile acids in the ileal digesta, while dietary DCA and commercial bile restored it. Together, this study showed that DCA and LCA reduced NE histopathology, suggesting that secondary bile acids, but not total bile acid levels, play an essential role in controlling the enteritis.

Duck viral enteritis is a highly contagious and fatal disease of commercial waterfowl flocks. The disease occurs sporadically or epizootically in mainland China due to insufficient vaccinations. Early and rapid diagnosis is important for preventive intervention and the control of epizootic events in clinical settings. In this study, we generated two monoclonal antibodies (MAbs) that specifically recognized the duck enteritis virus (DEV) envelope glycoprotein B and tegument protein UL47, respectively. Using these MAbs, a colloidal gold-based immunochromatographic assay (ICA) was developed for the efficient detection of DEV antigens within 15 min. Our results showed that the detection limit of the developed ICA strip was 2.52 × 103 TCID50/mL for the virus infected cell culture suspension with no cross-reactivity with other pathogenic viruses commonly encountered in commercially raised waterfowl. Using samples from experimentally infected ducks, we demonstrated that the ICA detected the virus in cloacal swab samples on day three post-infection, demonstrating an 80% concordance with the PCR. For tissue homogenates from ducks succumbing to infection, the detection sensitivity was 100%. The efficient and specific detection by this ICA test provides a valuable, convenient, easy to use and rapid diagnostic tool for DVE under both laboratory and field conditions.

Necrotic enteritis (NE) caused by C. perfringens is one of the most common diseases of poultry and results in a huge economic loss to the poultry industry, with resistant clostridial strains being a serious concern and making the treatment difficult. Whole-genome sequencing approaches represent a good tool to determine resistance profiles and also shed light for a better understanding of the pathogen. The aim of this study was to characterize, at the genomic level, a collection of 20 C. perfringens isolates from poultry affected by NE, giving special emphasis to resistance mechanisms and production of bacteriocins. Antimicrobial resistance genes were found, with the tet genes (associated with tetracycline resistance) being the most prevalent. Interestingly, two isolates carried the erm(T) gene associated with erythromycin resistance, which has only been reported in other Gram-positive bacteria. Twelve of the isolates were toxinotyped as type A and seven as type G. Other virulence factors encoding hyaluronases and sialidases were frequently detected, as well as different plasmids. Sequence types (ST) revealed a high variability of the isolates, finding new allelic combinations. Among the isolates, C. perfringens MLG7307 showed unique characteristics; it presented a toxin combination that made it impossible to toxinotype, and, despite being identified as C. perfringens, it lacked the housekeeping gene colA. Genes encoding bacteriocin BCN5 were found in five isolates even though no antimicrobial activity could be detected in those isolates. The bcn5 gene of three of our isolates was similar to one previously reported, showing two polymorphisms. Concluding, this study provides insights into the genomic characteristics of C. perfringens and a better understanding of this avian pathogen.

Bufavirus (BuV) and human parvovirus 4 (PARV4) belong to the Parvoviridae family. We assessed BuV and PARV4 DNA presence by real-time PCR analysis in stool, blood and respiratory samples collected in patients from Marseille and Nice, two large cities in the South-East of France. Bu-V DNA was detected in diarrheic stool samples from 92 patients (3.6% of 2583 patients), particularly men and adults, and patients from the nephrology and the infectious disease departments. Among the patients with a BuV-positive stool sample and for whom at least one blood sample was available (n = 30 patients), BuV DNA was detected also in 3 blood samples. In contrast, BuV DNA was not detected in any of the respiratory samples from 23 patients with BuV-positive stool. BuV detection rate was comparable in stool samples from patients with and without diarrhea. We did not detect PARV4 DNA in any of the stool specimens (n = 2583 patients). Our results suggest that PARV4 fecal-oral transmission is rare or non-existent in the South-East of France while BuV circulates with a relatively high rate in this area.

Objectives: To describe a case of Shanghai fever disease and to analyze other published reports in non-Asiatic countries, defining clinical characteristics and highlighting that this is not only an Asian disease. Study design: A computerized search without language restriction was conducted using PubMed and Scopus; all references listed were hand-searched to identify any other relevant literature. An article was considered eligible for inclusion in the systematic review if it reported cases with Shanghai fever described in non-Asiatic countries. Our case was also included in the analysis. Results: Ten articles reporting 10 cases of Shanghai fever disease were considered. Fever, diarrhea and ecthyma gangrenosum were the most frequent symptoms observed. Blood was the most common site of isolation for Pseudomonas aeruginosa. Three patients underwent surgery due to necrotizing enteritis and intestinal perforation. Meningitis was documented in one case. None of the patients received antipseudomonal antibiotics within 24 h of admission. The outcome was good in nine cases; only one patient died due to multiple organ failure from Pseudomonas sepsis. No common primary immune deficiency was identified in these patients. Extremely young age (<1 year) was the only host factor predisposing to Shanghai fever. Conclusions: It is important to shed light on this disease in non-Asiatic countries and take into account that it can also affect healthy children. Pediatricians, therefore, should consider Shanghai fever among diagnoses in children with community-onset diarrhea, fever and skin lesions suggestive of ecthyma gangrenosum to start an appropriate treatment sooner and to reduce the mortality in these children.

Infectious pancreatic necrosis (IPN), first described as acute viral catarrhal enteritis, is a highly contagious disease with variable pathogenicity that has been linked to genetic variation in the viral VP2 gene encoding the capsid protein. In this study, the IPN virus (IPNV) is isolated from the moribund fish from five of fourteen Iranian trout farms from 2015 to 2017. The affected fish showed mortality rates ranging from 20% to 60%, with the main clinical signs of exophthalmia, darkened skin, and mild abdominal distension, as well as yellow mucoid fluid in the intestine. Histopathological examination of intestinal sections confirmed acute catarrhal enteritis in all samples. RT-PCR assay of the kidney tissue and cell culture (CHSE-214) samples consistently confirmed the presence of the virus. The phylogenetic analysis of the partial VP2 sequence revealed that the detected isolates belong to genogroup 5, and are closely related to the Sp serotype strains of European origin. Characterization of VP2 of all isolates revealed the P217T221 motif that previously was associated with avirulence or low virulence, while all IPNV-positive fish in this study were clinically affected with moderate mortality. The IPNV isolates from Iran are associated with two lineages that appear to have originated from Europe, possibly via imported eggs.

Campylobacter spp. are major causes of foodborne illness globally, and are mostly transmitted through the consumption and handling of poultry. Campylobacter infections have widely variable outcomes, ranging from mild enteritis to severe illness, which are attributed to host interactions and the virulence of the infecting strain. In this study, in order to investigate the effect of host stress on the growth and pathogenicity of C. jejuni, three strains associated with human infection and two strains from broilers were subject to growth, motility, adhesion and invasion assays, in response to exposure to catecholamines; epinephrine, norepinephrine and the glucocorticoid neuroendocrine hormones corticosterone, cortisol and cortisone which are associated with stress in humans and broilers. Catecholamines resulted in significantly increased growth, adhesion and invasion of Caco-2 cells. Corticosterone promoted growth in one of five strains, and cortisone resulted in a significant increase in motility in two out of five strains, while no significant differences were observed with the addition of cortisol. It was concluded that stress-associated hormones, especially catecholamines, may promote growth and virulence in Campylobacter.

Clostridium perfringens is a versatile pathogen, inducing diseases in the skin, intestine (such as chicken necrotic enteritis (NE)), and other organs. The classical sign of NE is the foul smell gas in the ballooned small intestine. We hypothesized that deoxycholic acid (DCA) reduced NE by inhibiting C. perfringens virulence signaling pathways. To evaluate the hypothesis, C. perfringens strains CP1 and wild-type (WT) HN13 and its mutants were cultured with different bile acids, including DCA and isoallolithocholic acid (isoalloLCA). Growth, hydrogen sulfide (H2S) production, and virulence gene expression were measured. Notably, isoalloLCA was more potent in reducing growth, H2S production, and virulence gene expression in CP1 and WT HN13 compared to DCA, while other bile acids were less potent compared to DCA. Interestingly, there was a slightly different impact between DCA and isoalloLCA on the growth, H2S production, and virulence gene expression in the three HN13 mutants, suggesting possibly different signaling pathways modulated by the two bile acids. In conclusion, DCA and isoalloLCA reduced C. perfringens virulence by transcriptionally modulating the pathogen signaling pathways. The findings could be used to design new strategies to prevent and treat C. perfringens-induced diseases.

Chlamydia suis (C. suis) resides in the intestines of pigs and tetracycline-resistant strains are emerging worldwide. Intestinal infections are often subclinical. However, the gut is regarded as a C. suis reservoir and clinical infections have been associated with enteritis, conjunctivitis, pneumonia and reproductive failure. C. suis was found in boar semen and venereal transmission occurred. We studied the anti-Chlamydia suis activity of ovotransferrin (ovoTF) and bovine lactoferrin (bLF). Pre-incubation of C. suis with bLF or ovoTF had no significant effect on overall chlamydia replication (mean fluorescence area) in McCoy cells. The addition of ovoTF to the culture medium had no effect on bacterial replication, but the addition of 0.5 or 5 mg/mL of bLF significantly reduced the inclusion size by 17% and 15% respectively. Egg components are used for cryopreservation of boar semen. When inoculating an ovoTF-containing and Chlamydia suis-spiked semen sample in McCoy cells, a significant reduction in inclusion number (by 7%) and overall replication (by 11%) was observed. Thus, we showed that transferrins possess anti-chlamydial activity. Moreover, ovoTF addition to semen extenders might reduce C. suis venereal transmission. Further research is needed to unravel the mechanisms behind the observations and to enhance the effect of transferrins on C. suis.

Zoonotic Campylobacter, including C. jejuni and C. coli, are among the most prevalent agents of food-borne enteritis worldwide. The immunopathological sequelae of campylobacteriosis are caused by Toll-like Receptor-4 (TLR4)-dependent host immune responses, induced by bacterial lipooligosaccharide (LOS). In order to investigate C. coli-host interactions, including the roles of the human gut microbiota and TLR4, upon infection, we applied a clinical acute campylobacteriosis model, and subjected secondary abiotic, TLR4-deficient IL10-/- mice and IL10-/- controls to fecal microbiota transplantation derived from human donors by gavage, before peroral C. coli challenge. Until day 21 post-infection, C. coli could stably colonize the gastrointestinal tract of human microbiota-associated (hma) mice of either genotype. TLR4-deficient IL10-/- mice, however, displayed less severe clinical signs of infection, that were accompanied by less distinct apoptotic epithelial cell and innate as well as adaptive immune cell responses in the colon, as compared to IL10-/- counterparts. Furthermore, C. coli infected IL10-/-, as opposed to TLR4-deficient IL10-/-, mice displayed increased pro-inflammatory cytokine concentrations in intestinal and, strikingly, systemic compartments. We conclude that pathogenic LOS might play an important role in inducing TLR4-dependent host immune responses upon C. coli infection, which needs to be further addressed in more detail.

Human campylobacteriosis represents an infectious enteritis syndrome caused by Campylobacter species, mostly Campylobacter jejuni. Given that C. jejuni infections are rising worldwide and antibiotic treatment is usually not indicated, novel treatment options for campylobacteriosis are needed. Urolithin-A constitutes a metabolite produced by the human gut microbiota from ellagitannins and ellagic acids in berries and nuts which have been known for their health-beneficial including anti-inflammatory effects since centuries. Therefore, we investigated potential pathogen-lowering and immunomodulatory effects following oral application of synthetic urolithin-A during acute campylobacteriosis applying perorally C. jejuni infected, microbiota-depleted IL-10-/- mice as preclinical inflammation model. On day 6 post infection, urolithin-A treated mice harbored slightly lower pathogen loads in their ileum, but not colon as compared to placebo counterparts. Importantly, urolithin-A treatment resulted in an improved clinical outcome and less pronounced macroscopic and microscopic inflammatory sequelae of infection that were paralleled by less pronounced intestinal pro-inflammatory immune responses which could even be observed systemically. In conclusion, this preclinical murine intervention study provides first evidence that oral urolithin-A application is a promising treatment option for acute C. jejuni infection and paves the way for future clinical studies in human campylobacteriosis.

The European brown hare (Lepus europaeus) is a quite adaptable species, but populations have been decreasing for several decades in different countries, including Germany. To investigate infectious diseases as possible influences on observed population decline in the German federal state Schleswig-Holstein, 118 deceased free-ranging European brown hares were collected between 2017 and 2020 and underwent detailed postmortem examination with extended sampling. Infectious diseases were a major cause of death (34.7%). The number of juveniles found exceeded the adult ones. The main pathomorphological findings were hepatitis (32.8%), pneumonia (22.2%), nephritis (19.1%), liver necrosis (12.9%), and enteritis (40.7%). An unusual main finding was steatitis (20.9%) of unknown origin. Animals were mainly emaciated and showed high infection rates with Eimeria spp. (91.3%) and Trichostrongylus spp. (36.2%). European Brown Hare Syndrome Virus reached an epidemic status with few fatal infections (4.2%) and high seroprevalence (64.9%), whereas the prevalence of Rabbit Haemorrhagic Disease Virus 2 was very low (0.8%) in hares in Schleswig-Holstein. Pathogens such as Yersinia pseudotuberculosis (5.9%), Pasteurella multocida (0.8%), and Staphylococcus aureus (3.4%) only caused sporadic deaths. This study illustrates the wide distribution of various infectious pathogens with high mortality and even zoonotic potential. Infectious diseases need to be considered as an important influence on population dynamics in Schleswig-Holstein.

Chicken parvovirus (ChPV) is an agent frequently associated with runting stunting syndrome (RSS). This syndrome has been reported in association with ChPV in many countries, including Brazil; however, studies characterizing the virus on a molecular level are scarce, and ChPV pathogenicity in day-old chicks remains unclear. The aim of the present work was to establish the molecular characteristics of ChPV, determine the pathogenicity of ChPV in SPF chicks and detect and quantify ChPV by qPCR in several tissues and chicks of different ages. The experimental challenge was performed at one day of age, and daily and weekly observations were performed and five birds from each experimental group (mock and infected birds) were euthanized to perform the different analysis. ChPV genome copies were detected and quantified by qPCR in gut, spleen, thymus, kidney, pancreas, proventriculus and bursa. Clinically, the infected group presented with diarrhea 24 h post-infection, which persisted until 42 days of age. The small intestine was distended, and its contents were aqueous and foamy. Enteritis and dilated crypts with cyst shapes were observed in intestinal segments. Acute pancreatitis associated with lymphocytic nodules, infiltrating lymphocytes and plasma cells between the pancreatic acinus was observed. Koch's postulate was demonstrated and the genetic characterization of the VP1 gene showed that the Brazilian ChPV isolate belongs to the ChPV II group.

Campylobacter enteritis in humans is primarily associated with C. jejuni/coli infection. Other species cause campylobacteriosis relatively infrequently; while this could be attributed to bias in diagnostic methods, the pathogenicity of non-jejuni/coli Campylobacter spp. such as C. upsaliensis and C. helveticus (isolated from dogs and cats) is uncertain. Galleria mellonella larvae are suitable models of the mammalian innate immune system and have been applied to C. jejuni studies. This study compared the pathogenicity of C. jejuni, C. upsaliensis, and C. helveticus isolates. Larvae inoculated with either C. upsaliensis or C. helveticus showed significantly higher survival than those inoculated with C. jejuni. All three Campylobacter species induced indistinguishable histopathological changes in the larvae. C. jejuni could be isolated from inoculated larvae up to eight days post-inoculation whereas C. upsaliensis and C. helveticus could only be isolated in the first two days. There was a significant variation in the hazard rate between batches of larvae, in Campylobacter strains, and in biological replicates as random effects, and in species and bacterial dose as fixed effects. The Galleria model is applicable to other Campylobacter spp. as well as C. jejuni, but may be subject to significant variation with all Campylobacter species. While C. upsaliensis and C. helveticus cannot be considered non-pathogenic, they are significantly less pathogenic than C. jejuni.

Sepsis-associated acute liver injury caused by spillovers of bacteria and endotoxins (lipopolysaccharide, LPS) into the liver remains a public health issue due to the lack of specific therapeutic approaches. Previous studies showed that the recombinant protein HscB (rCsHscB) of Clonorchis sinensis, a carcinogenic liver fluke, had an anti-inflammatory effect and could alleviate inflammatory diseases such as enteritis; however, whether it can prevent sepsis-associated acute liver injury induced by LPS is still unknown. In our current study, the therapeutic effects and the potential mechanisms of rCsHscB on LPS-induced acute liver injury were investigated both in vivo and in vitro. The data showed that rCsHscB prevented LPS-induced liver damage, as demonstrated by histopathological observation and hepatic damage markers (the activities of serum ALT and AST) in a murine model of sepsis-associated acute liver injury. rCsHscB also significantly reversed the high levels of serum IL-6 and MCP-1 induced by LPS. In addition, rCsHscB attenuated the production of LPS-induced proinflammatory cytokines, including IL-6 and TNF-α, in a macrophage cell line-RAW264.7, through possible mediation by the MAPK signaling pathway in vitro. In conclusion, the present study demonstrates that rCsHscB derived from a fluke C. sinensis protects against sepsis-associated acute liver injury induced by LPS, which may be attributed to the inhibition of the MAPK signaling pathway. Our present study provides a potential therapeutic strategy for sepsis-associated acute liver injury.

Clostridium perfringens is the etiological agent for necrotic enteritis (NE) in broiler chickens, which causes a substantial economic loss of an estimated USD 6 billion annually in the global poultry industry. Collagen adhesion is involved in the NE pathogenesis in poultry. In this study, the binding capabilities of chicken C. perfringens isolates of various genetic backgrounds (netB-tpeL-, netB+tpeL-, netB+tpeL+) to collagen types I-V and gelatin were examined, and the putative adhesin protein cnaA gene was investigated at the genomic level. In total, 28 C. perfringens strains from healthy and NE-inflicted sick chickens were examined. The results on collagen adhesin-encoding gene cnaA by the quantitative-PCR results indicated that netB-tpeL- isolates had much lower copies of the detectable cnaA gene than netB+ isolates (10 netB+tpeL- isolates, 5 netB+tpeL+ isolates). Most of the virulent C. perfringens isolates demonstrated collagen-binding abilities to types I-II and IV-V, while some strains showed weak or no binding to collagen type III and gelatin. However, the netB+tpeL+ isolates showed significantly higher binding capabilities to collagen III than netB-tpeL- and netB+tpeL- isolates. The data in this study suggest that the collagen-binding capability of clinical C. perfringens isolates correlates well with their NE pathogenicity levels, especially for C. perfringens isolates carrying genes encoding crucial virulence factors and virulence-associated factors such as netB, cnaA, and tpeL. These results indicate that the presence of the cnaA gene may be correlated with C. perfringens virulence (particularly for netB+ isolates).

Campylobacter jejuni is a major foodborne pathogen and common cause of bacterial enteritis worldwide. A total of 622 C. jejuni isolates recovered from food animals and retail meats in the United States through the National Antimicrobial Resistance Monitoring System between 2013 and 2017 were sequenced using an Illumina MiSeq. Sequences were combined with WGS data of 222 human isolates downloaded from NCBI and analyzed by core genome multilocus sequence typing (cgMLST) and traditional MLST. cgMLST allelic difference (AD) thresholds of 0, 5, 10, 25, 50, 100 and 200 identified 828, 734, 652, 543, 422, 298 and 197 cgMLST types among the 844 isolates, respectively, and traditional MLST identified 174 ST. The cgMLST scheme allowing an AD of 200 (cgMLST200) revealed strong correlation with MLST. cgMLST200 showed 40.5% retail chicken isolates, 56.5% swine, 77.4% dairy cattle and 78.9% beef cattle isolates shared cgMLST sequence type with human isolates. All ST-8 had the same cgMLST200 type (cgMLST200-12) and 74.3% of ST-8 and 75% cgMLST200-12 were confirmed as sheep abortion virulence clones by PorA analysis. Twenty-nine acquired resistance genes, including 21 alleles of blaOXA, tetO, aph(3')-IIIa, ant(6)-Ia, aadE, aad9, aph(2')-Ig, aph(2')-Ih, sat4 plus mutations in gyrA, 23SrRNA and L22 were identified. Resistance genotypes were strongly linked with cgMLST200 type for certain groups including 12/12 cgMLST200-510 with the A103V substitution in L22 and 10/11 cgMLST200-608 with the T86I GyrA substitution associated with macrolide and quinolone resistance, respectively. In summary, the cgMLST200 threshold scheme combined with resistance genotype information could provide an excellent subtyping scheme for source attribution of human C. jejuni infections.