Prevalence of Arcobacter species on chicken carcasses during processing in Iran

The objective of this study was to determine the prevalence of Arcobacter spp. on chicken carcasses at different stages of broiler processing in a major commercial poultry processing plant in central Iran. Overall, 80 chicken carcasses were sampled from 5 sites along the processing line during a total of 10 visits. When the culture method was used, 185 of 400 (46.3%) carcasses were positive for Arcobacter. Arcobacter butzleri was more frequently isolated (82.7%) than Arcobacter cryaerophilus (12.4%) and Arcobacter skirrowii (4.9%). The frequency of Arcobacter spp. on carcasses was 36.3% before defeathering, 41.3% after defeathering, 48.8% after evisceration, 67.5% at 20 min postchilling, and 37.5% at 24 h postchilling. The frequency of Arcobacter spp.-positive carcasses was reduced in completely chilled chickens, but not during the slaughtering process. The PCR assay identified 57 Arcobacter-contaminated carcass samples that were negative when using the culture method. When the PCR method was used, the frequently of Arcobacter spp. on carcasses was 43.8% before defeathering, 45.0% after defeathering, 55.0% after evisceration, 88.8% at 20 min postchilling, and 85.7% at 24 h postchilling. Therefore, there was a high prevalence of Arcobacter spp., especially A. butzleri, in poultry carcasses. To our knowledge, the present study is the first report in which Arcobacter spp. were isolated from chicken carcasses in Iran.     

Occurrence and antimicrobial resistance of emergent Arcobacter spp. isolated from cattle and sheep in Iran

This study is conducted to determine the occurrence and antimicrobial resistance of Arcobacter spp. isolated from clinically healthy food animals. A total of 308 samples from cattle (200) and sheep (108) were collected from Shiraz slaughterhouse, southern Iran to investigate the presence of the important Arcobacter spp. using cultivation and Polymerase Chain Reaction (PCR) methods. Antimicrobial susceptibility of Arcobacter isolates was determined for 18 antibiotics using disk diffusion method. Among 308 samples, 27 (8.7%) and 44 (14.28%) were positive for the presence of Arcobacter species with cultivation and PCR procedures, respectively. The predominant species was A. butzleri in both cattle (58.33%) and sheep (55%). In addition, concurrent incidence of the species was observed in 25% of the positive samples. All Arcobacter isolates were resistant to rifampicin, vancomycin, ceftriaxone, trimethoprim and cephalothin. The isolates showed high susceptibility to tetracycline, oxytetracycline, erythromycin, ciprofloxacin, kanamycin, amikacin, gentamicin and enrofloxacin. No significant difference among cattle and sheep isolates in resistance pattern was observed. The results indicate that cattle and sheep are significant intestinal carriers for Arcobacter spp. Moreover, tetracycline and aminoglycosides showed great effects on Arcobacter species in antibiogram test and can be used for treatment of human Arcobacter infections.     

Detection of Helicobacter spp. in gastric, fecal and saliva samples from swine affected by gastric ulceration

The aim of this study was to evaluate the presence of Helicobacter (H.) spp. in swine affected by gastric ulceration. Stomachs from 400 regularly slaughtered swine were subjected to gross pathological examination to evaluate the presence of gastric ulcers. Sixty-five samples collected from ulcerated pars esophagea and 15 samples from non-ulcerated pyloric portions were submitted to histopathological and molecular analyses, to detect Helicobacter spp., H. suis and H. pylori by PCR. Feces and saliva swabs were also collected from 25 animals in order to detect in vivo the presence of Helicobacter spp.. Gastric ulcers were detected in 373 cases (93%). The presence of ulcers in association with inflammatory processes was further confirmed by histological examination. Forty-nine percent (32/65) of the ulcerated esophageal portions as well as 53% (8/15) of the non-ulcerated pyloric portions were positive for Helicobacter spp. by PCR. The Helicobacter spp. positive samples were also positive for H. suis, while H. pylori was not detected. These results were confirmed by restriction enzyme analysis. With regard to feces and saliva samples, 15/25 (60%) and 16/25 (64%) were positive for Helicobacter spp. PCR, respectively but all were negative in H. suis and H. pylori specific PCR.     

Diagnosis of Helicobacter spp. infection in canine stomach

A total of 75 biopsied samples of cardia, fundus, body, and pyloric antrum from necropsied dogs that were submitted to the Department of Pathology, Faculty of Veterinary Science, Chulalongkorn University from April 2003 to June 2004 were investigated. The objectives of this study were to determine the prevalence of Helicobacter spp. in canine stomach by polymerase chain reaction (PCR) in comparison to histochemistry versus immunohistochemistry (IHC), and to correlate these diagnostic methods with the clinical significance in infected dogs. Histopathological results revealed 60.0% (45/75) of samples to be positive, and consisted of mild gastritis in 64.44% (29/45), moderate gastritis in 11.11% (5/45), and severe gastritis in 24.44% (11/45). The proportion showing no histopathological lesions was 40.0% (30/75). Helicobacter spp. were localized to the luminal crypt in 18.67% (14/75), gastric pit in 22.67% (17/75), gastric gland in 21.33% (16/75), and gastric epithelium in 8% (6/75). The percentages of positive samples of Helicobacter spp. diagnosed by hematoxylin and eosin stain (H&E), Warthin Starry stain (WSS), IHC with rabbit polyclonal anti-H. pylori antibody, and PCR were 17.3% (13/75), 46.7% (35/75), 30.7% (23/75), and 10.7% (8/75), respectively. No significant differences weree observed in histopathological changes in portions of the stomach (p>0.05). The diagnosis of Helicobacter spp. by PCR in comparison to that by WSS and IHC was not significantly different (p>0.05). There were no relationships between pathological studies using H&E, WSS, and IHC, and especially between PCR and clinical signs of Helicobacter spp. infections in canine stomachs (p>0.05). The present study revealed significantly different levels of correlation for Helicobacter spp. detection between H&E and WSS (p<0.001). Results indicate that the method of choice for diagnosis of Helicobacter spp. infection in canine stomach is dependent on the purpose of study and appropriate specimen collection.     

Detection and identification of food‐borne pathogens of the genera Campylobacter,Arcobacter and Helicobacter by multiplex PCR in poultry and poultry products

The objective of this study was to develop a multiplex polymerase chain reaction (PCR) to detect and differentiate food‐borne pathogens of the three genera Campylobacter, Arcobacter and Helicobacter in a single step procedure. One common reverse primer and three genus‐specific forward primers were designed by hybridizing to the 16S rRNA of selected reference strains. Besides the species with significance as food‐borne pathogens isolated from poultry meat –Campylobacter jejuni, Campylobacter coli, Arcobacter butzleri and Helicobacter pullorum– several other members of these genera were tested to determine the specificity of the designed multiplex PCR. In total, 20 ATCC and NCTC reference strains of Campyobacter, Arcobacter and Helicobacter were used to evaluate the PCR. Specific amplificates were obtained from all thermophilic species of Campylobacter as well as from species of Arcobacter and Helicobacter. No amplification product was obtained from the non‐thermophilic Campylobacter, C. hyointestinalis and C. fetus. Furthermore, a total of 43 field strains of the three genera isolated from poultry, pigs, cattle and humans were investigated using this PCR. To confirm the classification of 10 H. pullorum strains the 16S rRNAs were sequenced. The developed PCR is a helpful diagnostic tool to detect and differentiate Campylobacter, Arcobacter and Helicobacter isolated from poultry and poultry products.     

Prevalence and antimicrobial resistance of Arcobacter species isolated from poultry meat in Iran

1. The objective of this study was to determine the prevalence and antimicrobial resistance of Arcobacter spp. isolated from different species of retail poultry meat in Iran.

2. From August 2012 to April 2013, a total of 540 raw poultry meat samples from chicken (n = 100), turkey (n = 100), quail (n = 100), partridge (n = 80), duck (n = 50), ostrich (n = 60) and geese (n = 50) were purchased from randomly selected retail outlets in Shahrekord, Isfahan, Sari and Rasht, Iran.

3. Using culture techniques, 71 of 540 poultry meat samples (13.1%) were positive for Arcobacter spp. The highest prevalence of Arcobacter spp. was found in chicken meat (28.0%), followed by quail (12.0%), duck (11.4%), turkey (11.0%), geese (8.0%), partridge (7.5%) and ostrich (3.3%) meat. The number of A. butzleri isolated from poultry meat samples (90.1%) was significantly higher than A. cryaerophilus (7.1%) and A. skirrowii (2.8%). Significantly more poultry meat samples were found to contain Arcobacter spp. by the PCR assay than by the culture method.

4. Susceptibilities of Arcobacter isolates were determined for 14 antimicrobial drugs using the disk diffusion method. All of the 71 Arcobacter isolates tested were resistant to one or more antimicrobial agents. Resistance to cephalothin and vancomycin (95.8%) was the most common finding, followed by resistance to methicillin, azithromycin and ampicillin. All Arcobacter isolates were susceptible to gentamicin, streptomycin, tetracyclin and kanamycin.

5. The results of this study indicated the importance of poultry meat, especially chicken meat, as potential sources of Arcobacter spp. infection in people. Furthermore, the strains indicated resistance to a broad spectrum of antibiotics.     

Identification of Helicobacter spp. in oral secretions vs. gastric mucosa of stray cats

The definite mode of transmission of Helicobacter infection is largely unknown. This study was carried out primarily, to determine the existence of Helicobacter spp. in the oral secretions of stray cats as one of the possible routes of transmission and secondly, to evaluate the accordance between oral and gastric colonization of Helicobacter spp. in these cats. Forty-three adult stray cats were thus studied for the presence of Helicobacter species by quantitative rapid urease test (RUT), cytology and PCR. Helicobacter spp. were found in the oral secretions and gastric biopsies of 93% and 67.5% of the stray cats, respectively. There was not, however, any agreement observed between Helicobacter colonization at these two locations, at neither genus nor species level. These findings suggest that the oral cavity is routinely exposed to transient forms of bacteria and may temporarily harbor Helicobacter spp. Thus, oral cavity as a source of Helicobacter spp. may act as a reservoir for transmission and may not necessarily reflect the colonization status of the gastric mucosa.     

Classification of Arcobacter species isolated from aborted pig fetuses and sows with reproductive problems in Brazil

Seventeen field isolates of Arcobacter species were recovered in Brazil from aborted porcine fetal livers (n = 3), kidneys (n = 2), and thoracic fluid (n = 1). Arcobacter species were also recovered from uterine and oviductal tissues (n = 5) and a placenta from sows with reproductive problems. These isolates were initially presumed to be Arcobacter cryaerophilus on the basis of aerobic growth at 30°C, indoxyl acetate hydrolysis, catalase and oxidase reactions, growth on MacConkey agar, sensitivity to 3.5% sodium chloride, and susceptibility to nalidixic acid (40 mg/ml). The isolates were confirmed as Arcobacter using polymerase chain reaction, and were classified as A. cryaerophilus 1A (24%), A. cryaerophilus 1B (71%), and A. butzleri (6%) using restriction fragment length polymorphism.     

Occurrence of Arcobacter in Iranian poultry and slaughterhouse samples implicates contamination by processing equipment and procedures

1. The occurrence of Arcobacter spp. and three pathogenic species of Arcobacter from Iranian poultry carcasses was investigated at different steps of broiler processing to determine critical control points for reducing carcass contamination.

2. Samples were collected from (a) cloaca immediately before processing, (b) different points during processing and (c) at different stations in a processing plant of a slaughterhouse in southern Iran.

3. After enrichment steps in Arcobacter selective broth, DNA of the samples was extracted and three significant pathogen species of Arcobacter were identified based on polymerase chain reaction (PCR) detection of 16S rRNA and specific species PCR.

4. Out of a total of 540 samples, 244 (45%) were positive for Arcobacter spp. Arcobacter butzleri was more frequently detected (73% ± 13.9%) than A. cryaeophilus (9% ± 13.9%) and A. skirrowii (4.1%). In addition, co-colonisation (A. butzleri and A. cryaerophilus) occurred in 13.9% of the positive samples.

5. The results indicate a high prevalence of Arcobacter in the investigated slaughterhouse and broiler carcasses and that Arcobacter is not a normal flora of the broilers. Evidence for the presence of Arcobacter in the environment and water of processing plants suggests that these are sources of contamination of poultry carcasses. In addition, contamination of the poultry carcasses can spread between poultry meats in different parts and processes of the slaughterhouse (pre-scalding to after evisceration).     

Survey of Helicobacter infection in domestic and feral cats in Korea

Discovery of Helicobacter (H.) pylori has led to a fundamental change in our understanding of gastric diseases in humans. Previous studies have found various Helicobacter spp. in dogs and cats, and pets have been questioned as a zoonotic carrier. The present study surveyed the Helicobacter infections and investigated the presence of H. felis and H. pylori infections in domestic and feral cats in Korea. Sixty-four domestic cats and 101 feral cats were selected from an animal shelter. Saliva and feces were evaluated by Helicobacter genus-specific polymerase chain reaction (PCR). Genus-specific PCR positive samples were further evaluated for H. felis and H. pylori using specific primer pairs. Thirty-six of 64 (56.3%) samples from domestic cats and 92 of 101 (91.1%) samples from feral cats were PCR positive; the positive rate of feces samples was higher than that of saliva samples in both groups. H. felis and H. pylori species-specific PCR was uniformly negative. The prevalence of Helicobacter spp. in feral cats was approximately two-fold higher than that of domestic cats. The fecal-oral route may be more a common transmission route not only between cats but also in humans.     

Occurrence of ɛ-proteobacterial species in rabbits (Oryctolagus cuniculus) reared in intensive and rural farms

In order to investigate the occurrence of Campylobacter, Helicobacter and Arcobacter species in caecal contents of rabbits reared in intensive and rural farms, a total of 87 samples from animals belonging to 29 farms were analysed by both cultural and PCR analyses.     

<Emphasis Type="Italic">Arcobacter</Emphasis> spp. in fecal samples from Brazilian farmed caimans (<Emphasis Type="Italic">Caiman yacare</Emphasis>, Daudin 1802)

The aim of this study was to perform the identification and molecular characterization of Arcobacter cryaerophilus and Arcobacter butzleri isolated from caiman (Caiman yacare), kept at a production farm, in Brazil. Forty fecal samples were analyzed. After isolation and identification, 21/40 strains of A. butzleri and 19/40 strains of A. cryaerophilus were subjected to PCR for potential virulence gene detection. The results of the PCR showed 38/40 strains positive for the cadF, cj1349, ciaB, and tlyA genes, 39/40 strains positive for the pldA gene, and 40/40 strains positive for the mviN gene. None of the strains presented the irgA gene. Hemagglutinin (hecA gene) and hemolysin (hecB) genes were detected in 21/40 and 16/40 strains, respectively. The SE-AFLP showed a great genetic diversity, but some clonally groups were disseminated in various tanks. These data reveal that the strains presented the same virulence traits described from Arcobacter isolated from food-borne disease in humans.     

A mixed population of Helicobacter pylori,Helicobacter bizzozeronii and “Helicobacter heilmannii” in the gastric mucosa of a domestic cat

Background

The presence of Helicobacter within the gastric mucosa is responsible for producing pathology in many animal species, including man. Since humans have been shown to harbour many of the same bacterial species as domestic carnivores, concern over their zoonotic potential has been growing. Helicobacter pylori, a class 1 carcinogen responsible for cases of gastritis and gastric cancer in humans, produces similar pathology in pet carnivores and is considered an example of anthroponosis. The case here presented refers to a 13 year-old mixed breed spayed female cat seen at necropsy.

Findings

Stomach samples were analysed for the presence of Helicobacter spp. by cytology, histopathology and PCR. Mild mucosal atrophy was observed in the fundus and antrum, while lymphoplasmocytic infiltrates where noted in the lamina propria of the antrum. Helicobacter-like organisms were observed in the corpus and antrum, occupying gastric glands and surface mucosa. It was possible to detect Helicobacter spp., H. pylori, H. heilmannii and H. bizzozeronii in the fundus, corpus and antrum by PCR, while in the antrum PCR samples were positive for H. pylori.

Conclusions

The spayed female under study could represent either a yet un-described population of domestic cats infected with H. pylori or a case of anthroponosis.     

The prevalence and anatomical distribution of equine gastric ulceration syndrome (EGUS) in 201 horses in Denmark

Reasons for performing study: The prevalence (up to 93% in Thoroughbred racehorses) and severity of equine gastric ulceration syndrome (EGUS) have been correlated with the type of training and associated management practices. However, there have been few reports to confirm these findings in nonracehorses in Europe. Objectives: To describe the prevalence, anatomical distribution, severity and number of gastric ulceration lesions in a population of Danish pleasure horses; and to investigate differences for groups based on age, breed type and workload. Methods: A total of 201 horses not in active race‐training, age 7 months‐27 years, were evaluated, representing 23 different stables from all 5 regions of Denmark. These horses were considered to be healthy by the owner and not on veterinary treatment for EGUS. Endoscopically observed ulcer lesion scores were based on the number present (0–4) and severity (0–5). The presence or absence of ulcers in the glandular and/or nonglandular regions of the stomach was recorded and which site the most severe ulcers were found. Results: The prevalence of EGUS severity score ≥2 was 53%. The most severe lesions were commonly observed at the margo plicatus. Although older horses were not more likely to be affected by clinically significant EGUS they were more likely to have lesions in both the glandular and nonglandular regions. Differences in location of EGUS lesions were identified in different age groups, breed types and in horses exposed to different levels of work. Conclusion and potential relevance: This study confirms that gastric ulceration can be prevalent in a group of apparently clinically normal horses, not in intensive work. Further investigation of reasons for differences in EGUS location between different populations may aid toward the development of novel preventive measures.     

Identification of Helicobacter DNA in feline pancreas,liver, stomach,and duodenum: Comparison between findings in fresh and formalin-fixed paraffin-embedded tissue samples

The clinical significance of Helicobacter spp. in feline digestive organs needs to be evaluated and formalin-fixed and paraffin-embedded (FFPE) tissue samples provide an invaluable source for molecular studies. In this study, we performed a PCR assay to investigate the presence of Helicobacter DNA in digestive organs from seven cats and compared this occurrence in fresh and formalin-fixed and paraffin-embedded (FFPE) tissue samples from the same organs. The present study identified Helicobacter DNA in the pancreas, liver, stomach, and duodenum in fresh tissue samples but only in the stomach in FFPE samples. To our knowledge this is the first time that Helicobacter DNA have been identified in the feline pancreas. This study indicates that it is important to be aware of differences between results when analyzing FFPE samples compared to fresh tissue samples, especially regarding longer DNA fragments (>200 bp (base pairs)).     

Isolation and preliminary characterization of a novel Helicobacter species from swine

OBJECTIVE: To determine whether a Helicobacter sp similar to Helicobacter pylori in the stomachs of humans could be isolated from the stomachs of pigs. ANIMALS: 4 young conventionally reared and 21 gnotobiotic pigs. PROCEDURE: Gastric mucosal homogenates (10% wt/vol) from 4 young conventionally reared pigs were cultured on Skirrow medium under microaerophilic conditions to assess the presence of Helicobacter spp. Colonies with morphologic features compatible with Helicobacter organisms were selected, tested for urease activity, and subpassaged on Skirrow medium. Isolates were examined via SDS-PAGE electrophoresis and reciprocal western blot analyses involving convalescent sera from monoinfected gnotobiotic pigs. RESULTS: Urease- and catalase-positive, gram-negative, microaerophilic, small, curved rod bacteria were isolated from the gastric mucosa of young healthy pigs. The first isolate (2662) was structurally and immunologically closely related to H pylori isolated from humans. The second isolate (1268) displayed an SDS-PAGE profile dissimilar to that of H pylori and isolate 2662, yet it shared limited immunologic cross-reactivity with these microbes. CONCLUSIONS AND CLINICAL RELEVANCE: Findings of this study indicate that development of gastric mucosal ulcers and ulceration of the nonglandular pars esophagea in pigs may be associated with gastric colonization by swine-origin Helicobacter spp, which are similar to H pylori isolated from humans.     

Helicobacter spp. in the saliva,stomach, duodenum and faeces of colony dogs

The role of Helicobacter spp. infection in canine gastrointestinal disease is unclear and routes of transmission are of epidemiological and zoonotic importance. The aim of this study was to identify Helicobacter spp. in the saliva, stomach, duodenum and faeces of dogs using a multiplex PCR, and to evaluate any attendant histopathological changes. Helicobacter canis was the most common species detected in saliva and faeces and no correlation between the presence of Helicobacter spp. and histopathological changes in either the stomach or duodenum was observed. All dogs examined were co-infected with up to four species of the organism. This is the first time these bacteria have been studied at species level at multiple sites within the canine alimentary tract.     

An investigation of Leishmania spp. in Didelphis spp. from urban and peri-urban areas in Bauru (São Paulo, Brazil)

Blood and bone marrow samples were taken from 112 Didelphis spp., collected between March 2005 and February 2006, from urban and peri-urban areas of Bauru, São Paulo State, Brazil, to evaluate the hypothesis that these animals might constitute a reservoir of Leishmania spp. Anti-Leishmania ssp. antibodies were screened in the serum samples using an enzyme-linked immuno-sorbent assay (ELISA) and the polymerase chain reaction (PCR). PCR was performed on fragments of DNA samples from Leishmania spp. using primers 13A and 13B, and showed a positive outcome in 91.6% of the 112 samples tested. Of the 107 samples analyzed by ELISA, 71% were positive. Evidence of epidemiological risk factors such as a circulating parasite and freely moving vectors suggests that Didelphis spp. may participate in the transmission cycle of Leishmania spp. in Bauru.     

Detection and characterization of Leptospira spp. in dogs diagnosed with kidney and/or liver disease in Selangor,Malaysia

Leptospirosis is a serious bacterial disease that affects both humans and animals. A wide range of symptoms have been described in humans; the disease in dogs is commonly associated with kidney and/or liver disease. In Malaysia, information about the common serovars infecting dogs is limited. Therefore, we investigated the occurrences of leptospirosis in 124 pet dogs diagnosed with kidney and/or liver disease. Blood, urine, abdominal effusion, and/or kidney and liver were collected from the dogs. Based on microscopic agglutination testing, 53 of 124 (42.7%) dogs were seropositive for leptospiral exposure. Sera were frequently positive to serovars Bataviae (n = 12), Javanica (n = 10), and Icterohaemorrhagiae (n = 10). Direct detection using PCR showed that 42 of 124 (33.9%) of the whole blood and 36 of 113 (31.9%) urine samples were positive for pathogenic Leptospira spp. By PCR, 2 of 23 (9.1%) kidney and 2 of 23 (9.1%) liver were positive for pathogenic Leptospira spp. Abdominal effusion from 4 dogs were PCR-positive for pathogenic Leptospira spp. The species detected were L. interrogans, L. borgpetersenii, L. kirschneri, and L. kmetyi by partial 16S rRNA sequencing. We further identified and characterized 11 Leptospira spp. isolates from 8 dogs as serovars Bataviae, Javanica, and Australis. The mortality rate of the Leptospira-infected dogs was high (18 of 53; 34%).     

Molecular biological identification of Babesia,Theileria, and Anaplasma species in cattle in Egypt using PCR assays,gene sequence analysis and a novel DNA microarray

In this preliminary study, a novel DNA microarray system was tested for the diagnosis of bovine piroplasmosis and anaplasmosis in comparison with microscopy and PCR assay results. In the Dakahlia Governorate, Egypt, 164 cattle were investigated for the presence of piroplasms and Anaplasma species. All investigated cattle were clinically examined. Blood samples were screened for the presence of blood parasites using microscopy and PCR assays. Seventy-one animals were acutely ill, whereas 93 were apparently healthy. In acutely ill cattle, Babesia/Theileria species (n = 11) and Anaplasma marginale (n = 10) were detected. Mixed infections with Babesia/Theileria spp. and A. marginale were present in two further cases. A. marginale infections were also detected in apparently healthy subjects (n = 23). The results of PCR assays were confirmed by DNA sequencing. All samples that were positive by PCR for Babesia/Theileria spp. gave also positive results in the microarray analysis. The microarray chips identified Babesia bovis (n = 12) and Babesia bigemina (n = 2). Cattle with babesiosis were likely to have hemoglobinuria and nervous signs when compared to those with anaplasmosis that frequently had bloody feces. We conclude that clinical examination in combination with microscopy are still very useful in diagnosing acute cases of babesiosis and anaplasmosis, but a combination of molecular biological diagnostic assays will detect even asymptomatic carriers. In perspective, parallel detection of Babesia/Theileria spp. and A. marginale infections using a single microarray system will be a valuable improvement.