The H9N2 avian influenza virus increases APEC adhesion to oviduct epithelia by viral NS1 protein-mediated activation of the TGF-ß pathway.

H9N2 avian influenza is a low-pathogenic avian influenza circulating in poultry and wild birds worldwide and frequently contributes to chicken salpingitis that is caused by avian pathogenic Escherichia coli (APEC), leading to huge economic losses and risks for food safety. Currently, how the H9N2 virus contributes to APEC infection and facilitates salpingitis remains elusive. In this study, in vitro chicken oviduct epithelial cell (COEC) model and in vivo studies were performed to investigate the role of H9N2 viruses on secondary APEC infection, and we identified that H9N2 virus enhances APEC infection both in vitro and in vivo. To understand the mechanisms behind this phenomenon, adhesive molecules on the cell surface facilitating APEC adhesion were checked, and we found that H9N2 virus could upregulate the expression of fibronectin, which promotes APEC adhesion onto COECs. We further investigated how fibronectin expression is regulated by H9N2 virus infection and revealed that transforming growth factor beta (TGF-ß) signaling pathway is activated by the NS1 protein of the virus, thus regulating the expression of adhesive molecules. These new findings revealed the role of H9N2 virus in salpingitis co-infected with APEC and discovered the molecular mechanisms by which the H9N2 virus facilitates APEC infection, offering new insights to the etiology of salpingitis with viral-bacterial co-infections.IMPORTANCEH9N2 avian influenza virus (AIV) widely infects poultry and is sporadically reported in human infections. The infection in birds frequently causes secondary bacterial infections, resulting in severe symptoms like pneumonia and salpingitis. Currently, the mechanism that influenza A virus contributes to secondary bacterial infection remains elusive. Here we discovered that H9N2 virus infection promotes APEC infection and further explored the underlying molecular mechanisms. We found that fibronectin protein on the cell surface is vital for APEC adhesion and also showed that H9N2 viral protein NS1 increased the expression of fibronectin by activating the TGF-ß signaling pathway. Our findings offer new information on how AIV infection promotes APEC secondary infection, providing potential targets for mitigating severe APEC infections induced by H9N2 avian influenza, and also give new insights on the mechanisms on how viruses promote secondary bacterial infections in animal and human diseases.

Gut microbiota and serum metabolome reveal the mechanism by which TCM polysaccharides alleviate salpingitis in laying hens challenged by bacteria.

This paper aimed to evaluate the effect of 3 kinds of TCM polysaccharides instead of antibiotics in preventing salpingitis in laying hens. After feeding the laying hens with Lotus leaf polysaccharide, Poria polysaccharide, and Epimedium polysaccharide, mixed bacteria (E. coli and Staphylococcus aureus) were used to infect the oviduct to establish an inflammation model. Changes in antioxidant, serum immunity, anti-inflammatory, gut microbiota, and serum metabolites were evaluated. The results showed that the 3 TCM polysaccharides could increase the expression of antioxidant markers SOD, GSH, and CAT, and reduce the accumulation of MDA in the liver; the contents of IgA and IgM in serum were increased. Decreased the mRNA expression of TLR4, NFκB, TNF-α, IFN-γ, IL1ß, IL6, and IL8, and increased the mRNA expression of anti-inflammatory factor IL5 in oviduct tissue. 16sRNA high-throughput sequencing revealed that the 3 TCM polysaccharides improved the intestinal flora disturbance caused by bacterial infection, increased the abundance of beneficial bacteria such as Bacteroides and Actinobacillus, and decreased the abundance of harmful bacteria such as Romboutsia, Turicibacter, and Streptococcus. Metabolomics showed that the 3 TCM polysaccharides could increase the content of metabolites such as 3-hydroxybutyric acid and isobutyl-L-carnitine, and these results could alleviate the further development of salpingitis. In conclusion, the present study has found that using TCM polysaccharides instead of antibiotics was a feasible way to prevent bacterial salpingitis in laying hens, which might make preventing this disease no longer an issue for breeding laying hens.

Lotus leaf extract can attenuate salpingitis in laying hens by inhibiting apoptosis.

This study aimed to determine whether the lotus leaf extract (LLE) had the effect of treating salpingitis in laying hens. First, the salpingitis model was established by the method of bacterial infection. Differential genes between salpingitis and healthy laying hens were identified by transcriptome sequencing, and GO and KEGG enrichment analyses were performed. Groups of treatment of antibiotics and LLE were established to verify the feasibility of the lotus leaf extract in treating salpingitis. Furthermore, the active component and pharmacological effects of LLE were identified using the UPLC-Q-TOF-MS and network pharmacology technique. At last, the mechanism of LLE treating salpingitis was further evaluated by DF-1 cells infected with bacteria. The results showed that LLE significantly reduced the levels of TLR4 and IFN-γ (P < 0.05), accelerated the levels of IgA and IgG (P < 0.05), regulated the levels of SOD and MDA (P < 0.05) in laying hens with salpingitis. A total of 1,874 differential genes were obtained according to the transcriptome sequencing. It was revealed a significant role in cell cycle and apoptosis by enrichment analysis. In addition, among the 28 components identified by UPLC-Q-TOF-MS, 20 components acted on 58 genes, including CDK1, BIRC5, and CA2 for treating salpingitis. After bacterial infection, cells were damaged and unable to complete the normal progression of the cell cycle, leading to cell cycle arrest and further apoptosis formation. However, with the intervention of LLE, bacterial infection was resisted. The cells proliferation was extensively restored, and the expression of NO was increased. The addition of LLE significantly decreased cell apoptosis. The G1 phase increased, the S phase and the G2 phase decreased in the model group; after the intervention of LLE, the G1 phase gradually returned to the average level, and G2 and S phases increased. The mRNA expression levels of BIRC5, CDK1, and CA2 were consistent with the predicted results in network pharmacology. At the same time, the mRNA expression levels of Caspase-3 and Caspase-7 were reduced after added with LLE. The mRNA expression levels of TNF-α, TRADD, FADD, Caspase-8, Caspase-10, and Caspase-9 (P < 0.05), which would inhibit death receptor activation and decrease the apoptotic cascade, were upregulated after bacterial infection. However, the results in LLE groups were downregulated (P < 0.05). Meanwhile, the mRNA expression levels of BCL-2 in LLE groups were increased significantly compared with it in model group (P < 0.05). Notably, LLE administration inhibited apoptosis and regulated the cell cycle distribution in the salpingitis induced by bacterial infection. These results indicated that the LLE attenuated bacterial-induced salpingitis by modulating apoptosis and immune function in laying hens.

Research Note: Disturbance of intracellular calcium signal in salpingitis simulation of laying hens.

This study investigated whether there is disturbance of calcium signal in the simulated salpingitis of laying hens. A total of 90 Roman Pink layers (81 wk; 1.916 ± 0.17 kg) were divided into 3 groups (Control treated with PBS, 1.85 mg lipopolysaccharide (LPS)/layer as LPS group, 1.85 mg LPS/layer as LPS+organic chemical reagent (OCR) group) with 6 replicates of 5 layers. Compared with the Control, the mRNA expression of calcium/calmodulin dependent protein kinase IV (CaMK IV), sarcoplasmic/endoplasmic reticulum calcium ATPase (SERCA), and plasma membrane calcium-transporting ATPase (PMCA) were not only decreased (P < 0.05) in magnum of laying hens from LPS and LPS+OCR groups, but also in isthmus and uterus of hens from LPS+OCR group. Moreover, the mRNA expression of calcium sensing receptor (CaSR) and Orai1 in uterus from LPS+OCR group were higher (P < 0.05) than that from Control. The relative fluorescence intensity of Ca2+ in uterus from LPS and LPS+OCR groups were significantly higher than that from Control (P < 0.05). In conclusion, it existed that the linkage of simulated salpingitis treated with LPS+OCR and altered intracellular calcium signals in layers, which provided a new insight for alleviating salpingitis and uterine dysfunction of laying hens.

End of lay postmortem findings in aviary housed laying hens.

Good health and low mortality are constitutive elements of good animal welfare. In laying hens, mortality and pathological findings are usually reported as cumulative proportions from onset of lay to culling. However, knowledge of pathological lesions and causes of death specifically toward the end of the production period are scarce. This study aimed to investigate the occurrence of postmortem lesions and tentative causes of death in non-beak trimmed, end of lay hens, housed in multitiered aviary systems. A convenience sample of 48 flocks was recruited. In each flock, layers dead between wk 65 and 70 were necropsied in the field. In total, 482 layers were subjected to postmortem examination. The 4 most common pathological lesions were keel bone fracture (KBF) (92%), fatty liver (42%), emaciation (23%), and salpingitis (22%). Apart from keel bone fracture, the relative frequency of the pathological lesions variated between flocks, indicating that flock is an important factor. Common tentative causes of death were salpingitis (18%) and fatty liver hemorrhagic syndrome (FLHS) (13%). This study sheds light on health challenges aviary housed layers are facing end of lay, which is crucial knowledge in the development of preventive measures to secure good health and welfare.

Effects of salpingitis simulation on the morphology and expression of inflammatory-related genes of oviduct in laying hens.

This study was conducted to simulate salpingitis of laying hens by observing the morphology and expression of inflammatory genes in the oviduct. A total of one hundred twenty 81-wk-old Roman Pink laying hens in good physical condition without the oviduct disease with an average egg production rate of 76% were fed a basal diet for 2 wks and then randomly allocated into 4 groups (6 replicates/group, 5 birds/replicate). The experimental treatments were as follows: 1) Control group (treated with PBS); 2) Organic chemical reagent (OCR) group; 3) Lipopolysaccharide (LPS) group; 4) LPS + OCR group. First, the chickens were kept upside down to make ectropion and exposure of the apertura uterinae; then prepared reagents were poured into the uterine part of the fallopian tube by using the chicken vas deferens (1 mL/layer); finally, the chickens were kept in the inverted position for 5 to 10 min. The fallopian tube samples (the magnum, isthmus, and uterus) were collected after 48 h of treatment. Compared with the control, treatment with LPS+OCR decreased (P < 0.05) the secondary villus length and primary villus area in magnum and villus length in isthmus (P < 0.05). An increase (P < 0.05) of the intervillous space of uterus was observed in LPS + OCR group compared with the control. The expressions of interleukin-6 mRNA of magnum and interferon-γ (IFN-γ) of isthmus in the LPS and LPS+OCR treatments were higher (P < 0.05) than that in control. Compared with the control, treatment with LPS+OCR increased (P < 0.05) the expressions of IFN-γ mRNA of magnum and IFN-γ, tumor necrosis factor-α and inducible nitric oxide synthase mRNA of uterus in laying hens. In conclusion, the results of morphological damage of fallopian tube tissue and increased expression of inflammatory factors in LPS + OCR treatment group suggested that LPS+OCR treatment can provide data basis to establish salpingitis model in laying hens for studying the pathogenesis of it.

Bovine salpingitis: Histopathology, bacteriology, cytology and transcriptomic approaches and its impact on the oocyte competence.

The present study was performed to examine the histopathology, cytology, bacteriology and expression pattern of a targeted set of genes of cytokines in the oviduct of cows with inflammation (Experiment 1). In addition, the effects of oviductal fluid from cows with salpingitis on the oocyte maturation and fertilization in vitro were examined (Experiment 2). The most frequent bacterial co-infection was Escherichia coli and Fusobacterium necrophorum, which was always associated with severe histopathologic salpingitis. Out of 15 cows with histologically healthy uterus, only one cow (6.7%) displayed the histologic signs of mild salpingitis, whereas from 50 cows with endometritis, 48 cows (96%) showed histologically different grades of salpingitis. The mRNA expression of IL1ß, CD14, IL8 and CASP3 was significantly different among all groups of salpingitis (P < 0.05) with the highest level of mRNA expression in the sever grade of salpingitis. Results of experiment 2 showed a significant decline in the oocytes with peripheral free mitochondria and fertilization rate in the salpingitis group than the no- salpingitis group (P < 0.05). In conclusion, our results showed that histologically detected salpingitis is in most cases associated with histologic and cytologic endometritis. The pattern of the gene expression of chemokines and cytokines was altered in association with different grades of salpingitis. Further, we observed a decline in the peripherally located mitochondria and lower fertilization rate in oocytes following addition of oviductal fluid collected from the cows with sapingitis to the maturation media.

Necrotizing Salpingitis by Fowl Adenovirus in a Backyard Hen.

A 3-yr-old Ameraucana hen was received for postmortem examination following a 1-day history of lethargy and death. Gross lesions observed during necropsy were limited to pulmonary congestion and a small clump of egg yolk material in the oviductal lumen. On histopathology, there was a necrotizing salpingitis of the infundibular and isthmus mucosa with amphophilic, intranuclear inclusion bodies in superficial epithelial cells. Transmission electron microscopy identified the intranuclear inclusions as aggregates of adenovirus virions. Fowl adenovirus (FAdV) type A was identified with PCR and sequencing. Although the cause of death was not determined in this case, this is the first report of FAdV type A-associated salpingitis in a hen.

Reporte de caso- Salpingitis necrotizante por adenovirus en una gallina de traspatio. Una gallina de tres años fue recibida para examen post-mortem después de sufrir letargia por un día y la muerte. Las lesiones macroscópicas observadas durante la necropsia se limitaron a congestión pulmonar y pequeñas cantidades de yema de huevo en el lumen del oviducto. A través del examen histopatológico se observó una salpingitis necrotizante en la mucosa del infundíbulo e istmo con cuerpos de inclusión intranucleares y anfofílicos en las células epiteliales superficiales. Con el uso de microscopía electrónica de transmisión se determinó que las inclusiones intranucleares consistían en agregados de viriones de adenovirus. Se identificó adenovirus del pollo tipo A (FAdV) mediante PCR y secuenciación. Aunque la causa de muerte no fue determinada en este caso, este es el primer reporte de salpingitis asociada a la infección por adenovirus del pollo tipo A en una gallina.

Whole genome sequence comparison of avian pathogenic Escherichia coli from acute and chronic salpingitis of egg laying hens.

BACKGROUND: Infection in the oviduct (salpingitis) is the most common bacterial infection in egg laying hens and is mainly caused by Escherichia coli. The disease is responsible for decreased animal welfare, considerable economic loss as well as a risk of horizontal and vertical transmission of pathogenic E. coli. The outcome of salpingitis may be either acute or chronic. It has not yet been clarified whether the pathological manifestation is a result of the characteristics of the E. coli or whether the manifestation is associated with host factors such as host immunity. RESULTS: From the core- and accessory genome analysis and comparison of 62 E. coli no genetic markers were found to be associated to either acute or chronic infection. Twenty of the 62 genomes harboured at least one antimicrobial resistance gene with resistance against sulfonamides being the most common. The increased serum survival and iron chelating genes iss and iroN were highly prevalent in genomes from both acute and chronic salpingitis. CONCLUSION: Our analysis revealed that no genetic markers could differentiate the E. coli isolated from acute versus chronic salpingitis in egg laying hens. The difference in pathological outcome may be related to other factors such as immunological status, genetics and health of the host. These data indicate that salpingitis is another manifestation of colibacillosis.

Virulence of Escherichia coli Isolates Obtained from Layer Chickens with Colibacillosis Associated with Pericarditis, Perihepatitis, and Salpingitis in Experimentally Infected Chicks and Embryonated Eggs.

To evaluate the virulence of avian pathogenic Escherichia coli (APEC) isolates obtained from colibacillosis cases associated with pericarditis, perihepatitis, and salpingitis, the embryo lethality assay and experimental infection model in chicks were used in this study. According to the established criteria based on mortality in the embryo lethality assay for evaluating the virulence of E. coli isolates, 23 of the 26 APEC isolates associated with pericarditis and perihepatitis and 8 of the 20 isolates associated with salpingitis were found to be virulent. Isolate D137, which had been obtained from a case with pericarditis and perihepatitis and had an embryo mortality of 92%, and isolate D445, which had been obtained from a case with pericarditis and perihepatitis and had an embryo mortality of 17%, were used for the experimental infection. Four of the five 11-day-old chickens inoculated through the air sac with isolate D137 died 1 day postinoculation, and the challenge strain was recovered from the air sac, pericardial sac, or liver; however, colibacillosis lesions were found in only one of the five birds postmortem. All five chicks inoculated with isolate D445 survived for 7 days postinoculation and exhibited airsacculitis or pericarditis lesions at 7 days postinoculation; the challenge strain was not recovered from the lesions postmortem. The results obtained in this study suggest that the different APEC isolates tested cause illness in chickens through distinct pathogenesis.

Experimental induced avian E. coli salpingitis: Significant impact of strain and host factors on the clinical and pathological outcome.

Several types of Escherichia coli have been associated with extra-intestinal infections in poultry, however, they may vary significantly in their virulence potential. The aim of the present study was to investigate the virulence of five strains of E. coli obtained from different disease manifestations or from the cloacae of a healthy chicken. The virulence potential of the strains were evaluated in an avian experimental model for ascending infections, and experiments were conducted in both layers and broiler breeders. The clinical outcome of infection was highly depending on the challenge strain, however, not significantly reflecting the origin of the strain. In general, broiler breeders had a more severe clinical outcomes of infection compared to layers, but major with-in group diversity was observed for all challenge strains of clinical origin. A single strain of ST95 (phylogroup B2) had a distinct ability to cause disease. Results of the study shows major differences in virulence of different strains of E. coli in ascending infections; however, there was no indication of tissue-specific adaptation, since strains obtained from lesions unrelated to the reproductive system were fully capable of causing experimental infection. In conclusion, the study provides evidence for the clinical outcome of infection with E. coli in poultry is largely influenced by the specific strain as well as individual host factors.

Pathology and Molecular Characterization of Escherichia Coli Associated With the Avian Salpingitis-Peritonitis Disease Syndrome.

Outbreaks of salpingitis and peritonitis cause major economic losses due to high mortality, reduced egg-production, and culling. The aim of the present study was to characterize, in detail, lesions associated with increased mortality in layers due to avianpathogenic Escherichia coli (APEC) and to investigate the population structure of the E. coli involved, which is important for selection of optimal treatment and prophylactic strategies. Among 322 layers received from eight farms with increased mortality due to E. coli, three lesion types were observed; sepsis-like lesions, chronic salpingitis and peritonitis, and chronic salpingitis and peritonitis associated with sepsis-like lesions. One hundred isolates of E. coli obtained in pure culture from the different lesion types were selected for genetic characterization. Six out of 10 submissions (two farms with two submissions) were considered clonal as defined by more than 85% of the typed isolates of E. coli belonging to the same sequence-type (ST). B2 was the most-prevalent phylogroup, including the clonal complex of ST95. The most-important virulence genes of E. coli were demonstrated from both clonal and nonclonal outbreaks, and major differences as to phylogeny and virulence genes were not observed between the lesion types. Cannibalism was more-often observed during polyclonal outbreaks. A new pathotype of APEC is suggested based upon lesions and route of infection, high similarity of virulence genes including plasmid-associated genes, and high frequency of ST95 and other isolates belonging to phylogroup B2. Compared to the best-known pathotypes of E. coli, this needs further investigations, including infection experiments to show if single virulence factors can be pointed out that are specific for the salpingitis-peritonitis pathotype and possibly not found in other pathotypes of E. coli.

Occurrence of weak mutators among avian pathogenic Escherichia coli (APEC) isolates causing salpingitis and peritonitis in broiler breeders.

A collection of 46 avian pathogenic Escherichia coli (APEC) isolates was examined for the presence of mutators by determining the rate of mutation to rifampicin resistance. The collection included 34 E. coli isolates obtained in pure culture from chronic lesions of salpingitis and peritonitis in 34 broiler breeders, of which 12 were associated with the development of secondary septicemia. Twelve additional isolates were obtained from a clonal outbreak (ST95) of E. coli peritonitis syndrome (EPS), the lesions of which changed gradually over time into a subacute/chronic form. The hypothesis of the present study was that mutation rates would be higher for chronic infection isolates than for isolates from acute infections/exacerbations. The distribution of mutation rates followed a pattern similar to that found for other clinical isolates of E. coli, with a modal/median value of 1.47 × 10(-8). Of the 46 isolates, 24% (n=11) were weakly hypermutable (2.00 × 10(-8) ≤ µ<2.00 × 10(-7)), however, no strong mutators were detected (µ ≥ 2.00 × 10(-7)). Chronic salpingitis isolates had the highest proportion (45%, P=0.001) of weak mutators and also, significantly higher mutation rates (P=0.003) compared to isolates that caused septicemia (4%). In addition, mutation rates were significantly lower among ST95 isolates (P<0.0005), and among isolates from the same clonal group as ST95 (P=0.027), when compared to isolates from other groups. Although a clear association with the time phase of infection (as lesions of EPS became more chronic) could not be observed (ρ=0.523, P=0.081), a higher frequency of weak mutators among chronic infection isolates suggests that increased mutation rates play a role in adaptation of APEC to long-term persistence in an infected host environment.

Human and avian extraintestinal pathogenic Escherichia coli: infections, zoonotic risks, and antibiotic resistance trends.

Extraintestinal pathogenic Escherichia coli (ExPEC) constitutes ongoing health concerns for women, newborns, elderly, and immunocompromised individuals due to increased numbers of urinary tract infections (UTIs), newborn meningitis, abdominal sepsis, and septicemia. E. coli remains the leading cause of UTIs, with recent investigations reporting the emergence of E. coli as the predominant cause of nosocomial and neonatal sepsis infections. This shift from the traditional Gram-positive bacterial causes of nosocomial and neonatal sepsis infections could be attributed to the use of intrapartum chemoprophylaxis against Gram-positive bacteria and the appearance of antibiotic (ATB) resistance in E. coli. While ExPEC strains cause significant healthcare concerns, these bacteria also infect chickens and cause the poultry industry economic losses due to costs of containment, mortality, and disposal of carcasses. To circumvent ExPEC-related costs, ATBs are commonly used in the poultry industry to prevent/treat microbial infections and promote growth and performance. In an unfortunate linkage, chicken products are suspected to be a source of foodborne ExPEC infections and ATB resistance in humans. Therefore, the emergence of multidrug resistance (MDR) (resistance to three or more classes of antimicrobial agents) among avian E. coli has created major economic and health concerns, affecting both human healthcare and poultry industries. Increased numbers of immunocompromised individuals, including the elderly, coupled with MDR among ExPEC strains, will continue to challenge the treatment of ExPEC infections and likely lead to increased treatment costs. With ongoing complications due to emerging ATB resistance, novel treatment strategies are necessary to control ExPEC infections. Recognizing and treating the zoonotic risk posed by ExPEC would greatly enhance food safety and positively impact human health.

Genetic diversity and virulence profiles of Escherichia coli causing salpingitis and peritonitis in broiler breeders.

The genetic relatedness and virulence profiles of avian pathogenic Escherichia coli that caused salpingitis and peritonitis in 68 broiler breeders from 21 Danish farms were determined by multilocus sequence typing (MLST), pulsed-field-gel-electrophoresis (PFGE), ECOR phylogrouping, and PCR-based virulotyping. Phylogroups A, B1, B2, and D accounted for 19.1%, 5.9%, 52.9%, and 22.1% of the isolates, respectively. Overall, a total of five main MLST-based clonal groups (3-38 isolates) were identified, comprising 85.3% of the isolates. The most common sequence type (ST) was ST95 (n=12), followed by the ST428-, ST23- and ST350-clonal complexes (CCs) (n=8, n=7 and n=6, respectively). The emerging, antimicrobial resistance-associated clones, ST131 and ST648, were represented by five and three isolates, respectively, whereas ST352 and the ST168 CC comprised four isolates each. Phylogroup-B2 isolates showed a greater prevalence of nine virulence genes (P<0.05). One specific clonal group was significantly associated with phylogroup-B2 isolates (P<0.001), and with isolates that induced secondary septicemia (P=0.001). PFGE analysis revealed 12 clusters of genetically related strains (2-12) sampled from unrelated and geographically distant farms, indicating the widespread distribution and recent vertical transmission of particular APEC lineages. Certain lineages showed more diversity, substantiating that long-term, endemic transmission has been maintained. In conclusion, endemic lineages of E. coli that cause salpingitis and peritonitis in broiler breeders, although diverse, tend to be phylogenetically related, and demonstrate conserved virulence genotypes that might be associated with greater pathogenic potential.

Salpingitis in geese associated with Mycoplasma sp. strain 1220.

An outbreak of disease in a White Rhine laying goose flock was characterized by increased water uptake, increased mortality, production of eggs with abnormal shells, a 25% drop in egg production and 40% embryo mortality. Affected dead or sacrificed birds had sero-fibrinogranulocytic peritonitis and salpingitis, infiltration of the lamina propria in the uterus and heterophil granulocytes in the isthmus and magnum of the oviduct. Mycoplasmas, mainly identified as Mycoplasma sp. strain 1220, were isolated from the airsac, liver, ovary, magnum and peritoneum of some affected geese. Strain 1220 was originally isolated from a Hungarian gander with phallus inflammation and, according to detailed biochemical and serological examinations, it is expected to represent a new avian species within the genus Mycoplasma.

Multiple routes of entry for Escherichia coli causing colibacillosis in commercial layer chickens.

Colibacillosis associated with salpingitis occurred in a layer chicken flock on a commercial egg-producing farm in Japan. An increase in mortality was observed when the birds were at 62 weeks of age and reached 0.89% at 68 weeks of age. Postmortem examinations revealed pericarditis, perihepatitis, airsacculitis, and reproductive tract lesions in 4 affected birds at 69 weeks of age. Analysis of pulse-field gel electrophoresis (PFGE) patterns and putative virulence genes of 22 E. coli isolates obtained from the affected birds demonstrated that isolates from liver, heart, and the surface of the reproductive tract of one bird were genetically unrelated with those recovered from the lumen of the oviduct. In the other birds, isolates from liver, heart, and reproductive tract lesions were closely related to each other. These findings suggest that salpingitis in the former bird may be caused by ascending infection of the oviduct from the cloaca and salpingitis in the remaining birds may occur as part of systemic infection.

Molecular epidemiology of a reproductive tract-associated colibacillosis outbreak in a layer breeder flock associated with atypical avian pathogenic Escherichia coli.

The molecular epidemiology of 70 Escherichia coli isolates from an infection outbreak in a layer breeder flock was examined by pulsed-field gel electrophoresis and for a range of virulence factors by polymerase chain reaction. Pulsed-field gel electrophoresis showed 35 of 45 isolates from eight disease cases were associated with a single clonal group that was the exclusive strain associated with reproductive tract. A second unrelated group was found in environmental isolates and healthy birds. The remaining isolates were unrelated to each other or either clonal group. Polymerase chain reaction virulotyping indicated the "epidemic" clonal group contains virulence factors including iss, sfa, tsh, iucC, ibeA, and sitA associated with avian pathogenic E. coli plus several virulence factors more normally associated with human urinary tract infection. Significantly, the "epidemic" clone was also found in an environmental sample, suggesting it may have been transmitted to the flock via the environment.

[Escherichia coli salpingitis and peritonitis in layer chickens: an overview]. / Escherichia coli-salpingitis en -peritonitis bij leghennen: een overzicht.

Escherichia coli can induce salpingitis and/or peritonitis, a major cause of mortality in layer hens, but also other localized and systemic infections. E. coli infections have also been described in turkeys, geese, and ducks and are thought to be the cause of significant economic losses. However little is known about the real economic impact of the disease in layer chickens. The pathogenesis of E. coli salpingitis and peritonitis has not been elucidated yet. Three routes of infection have been discussed in the literature: ascending faecal contamination from the cloaca, bacterial translocation from the respiratory tract (air sac and lungs) and bacterial translocation from the intestinal lumen. Only one study has reported the occurrence of ascending faecal contamination from the cloaca to the oviduct and subsequently to the peritoneum. Regarding bacterial translocation, the only models available are for mammals, and these have not been applied to chickens so far Animal models could prove valuable to elucidate the pathogenesis of E. coli-induced salpingitis and peritonitis, and for assessing the value of preventive and curative intervention strategies. Little is known about risk factors for E. coli salpingitis and peritonitis. In contrast to colibacillosis in broilers, recent research has failed to demonstrate an association between several pathogens of the respiratory tract and the occurrence of E. coli pathology in layer chickens. The distance between poultry farms and the hen density in the cages were recently proposed as important risk factors for outbreaks ofcolibacillosis in flocks of layer hens, while in the past hormonal factors were implicated. The latter is an area of research that deserves more attention. Several methods for the molecular typing of E. coli have been described and might prove useful to study the epidemiology ofE. coli outbreaks in poultry, about which little is known. The presumptive diagnosis E. coli salpingitis and peritonitis is rather simple to establish, based on the anamnesis, clinical symptoms, and macroscopic findings at post-mortem. However; bacteriological analysis is required to establish a definite diagnosis because other pathogens can also cause salpingitis and peritonitis in layer hens. Antibiotics, chosen on the basis of sensitivity testing and their pharmacokinetic properties can be used as therapy; however residues in eggs may occur. Autovaccines are often used as prevention because in practice effective protection is only achieved against homologous E. coli serotypes.

Observations on salpingitis, peritonitis and salpingoperitonitis in a layer breeder flock.

A flock of 13,951 hens and 1379 cockerels was monitored from 26 to 58 weeks of age for the complex of salpingitis, peritonitis and salpingoperitonitis (sps). Two hundred and forty-three hens (78 per cent of the hens that died) were examined postmortem, and sps was recognised by gross examination for inflammatory exudate, in the body cavity or oviduct in 111 (46 per cent) of them. Salpingoperitonitis was the most common form, followed by salpingitis and then peritonitis. There were acute and chronic cases in all three conditions, but only in peritonitis were acute cases more common than chronic cases. Seventeen birds that had died of sps were cultured for aerobic bacteria within 12 hours of death. Escherichia coli was recovered from a variety of tissues from all of them, and other bacteria, including staphylococci, Mannheimia haemolytica and Streptococcus bovis, were isolated from a few carcases, either alone or together with E coli. Relatively few isolations of E coli were made from normal hens cultured 48, 72 and 96 hours after death.