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Acta Veterinaria Hungarica
Authors:
Simonetta Appino
,
F. Guarda
,
Paola Pregel
,
S. Amedeo
,
M. A. Cutufia
,
Giuseppina Bellonio
, and
A. Ponzetto

The aim of this study was to evaluate by PCR the presence of Helicobacter spp. in gastric mucus from the fundic region of the stomach and to investigate its role in oesophagogastric ulcers in swine bred and regularly slaughtered in Piedmont (Northern Italy). Stomachs from 595 regularly slaughtered swine were subjected to gross pathological examination in order to evaluate the presence of gastric ulcers (revealed in 75 cases, 12.6%). Histopathological examination was performed to better characterise erosions and ulcers. DNA extracted from gastric mucus collected from all the ulcer-affected and from 25 normal stomachs was submitted to PCR using Helicobacter spp. 16S rRNA gene target primers. Sixty-three percent (47/75) of the affected stomachs was positive as well as 24% (6/25) of the non-affected ones. Sequence analysis from 5 positive samples showed 99% homology with Helicobacter candidatus suis 16S ribosomal RNA gene.

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European Journal of Microbiology and Immunology
Authors:
Alexis Lacout
,
Marie Mas
,
Julie Pajaud
,
Véronique Perronne
,
Yannick Lequette
,
Michel Franck
, and
Christian Perronne

patients suffering from polymorphic signs and symptoms (SPPT/PTLDS), using real time qPCR, which is a direct diagnostic method amplifying the DNA of the microorganisms sought. As the yield of PCRs looking for Borrelia or co-infections in the venous blood is

Open access

. Pierce VM , Elkan M , Leet M , McGowan KL , Hodinka RL : Comparison of the Idaho Technology FilmArray system to real-time PCR for detection of respiratory pathogens in children . J Clin Microbiol 50 , 364 – 371 ( 2012

Open access

Adenoviruses are frequent infectious agents in different poultry species. The traditional, serological typing of new isolates by virus neutralisation tests is now in transition to be replaced by PCR and sequencing. The first PCRs, recommended for the detection of adenoviruses, had been designed to target the gene of the major capsid protein, the hexon. In birds, members of three different genera of the family Adenoviridae may occur. Accordingly, three specific hexon PCRs had to be elaborated for the detection of adenoviruses in poultry. A significantly more sensitive PCR, targeting the viral DNA-dependent DNA polymerase gene, has been described recently. This method proved to be an efficient alternative for the general detection of adenoviruses irrespective of their genus affiliation. Fowl adenoviruses (FAdVs), isolated from chicken to date, comprise twelve serotypes classified into five virus species (FAdV-A to E). The polymerase gene sequence has been determined yet only from three FAdV types representing three species. In the present work, the panel of polymerase gene sequences was completed with those of the rest of FAdVs. The newly determined sequences will facilitate the identification of new FAdV isolates as an existing species or as a putative new FAdV. Once the polymerase sequence is known, more specific PCRs for the amplification of the hexon and other genes can be designed and performed according to the preliminary species classification.

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Acta Veterinaria Hungarica
Authors:
HongBin Yan
,
XinWen Bo
,
Youyu Liu
,
Zhongzi Lou
,
XingWei Ni
,
WanGui Shi
,
Fang Zhan
,
HongKean Ooi
, and
WanZhong Jia

Moniezia benedeni and M. expansa are common ruminant tapeworms of worldwide distribution, causing gastrointestinal disorders and even death in sheep and goats. In this study, a polymerase chain reaction- (PCR-) based approach for precise species identification was developed and also applied to faecal DNA diagnosis of the tapeworm infection. Since nuclear ribosomal DNA (rDNA) appears to be a useful target for species and/or strain markers, the 18S regions of the rDNA of M. benedeni and M. expansa were amplified and sequenced. The lengths and GC contents of the regions sequenced were 2476–2487 bp and 51.9–52.1% for M. benedeni and 2473 bp and 51.9–52.0% for M. expansa, respectively. Alignment and comparison of the 18S sequences of the two species showed 92.5–93.3% homology. No matches for the 18S regions of M. benedeni and M. expansa were found with other species by BLAST search, which made the 18S sequences appropriate markers for the design of distinctive primers for the two Moniezia species. Our 18S-based PCR could detect as low levels as 0.5 pg genomic DNA or the DNA extracted from 0.2 g faecal sample collected from the rectum of an M. expansa-infected goat. The results indicate that this PCR approach is a reliable alternative for the differential diagnosis of Moniezia species in faecal samples.

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Analysis of flagellin genes was carried out on strains of Salmonella Typhimurium, Salmonella Hadar, Salmonella Abortusequi, Salmonella Enteritidis and Salmonella Gallinarum serovars, using a PCR system designed in this study. The purpose of these studies was to explore the flagellin genes of biphasic and monophasic Salmonellae for future targeted genetic interventions. The PCR primers were designed for two different structural genes of flagellin (fliC, fljB), for the repressor of fliC (fljA), for the operator region of fliC, and for the invertase system responsible for phase variation in Salmonella (hin, hixL, hixR). PCR analysis revealed that all of the examined genes (fliC, fliC-operator, fljB, fljA, hin, hixL, hixR) were present in all S. Typhimurium (n = 10)and S. Hadar (n = 10) strains tested. The results obtained on S. Typhimurium and S. Hadar strains confirmed their biphasic character at DNA level. However, the S. Enteritidis (n = 46) and S. Gallinarum (n = 5) strains lacked the invertase system (hin, hixL, hixR) as well as the fljA and fljB genes, while fliC and its operator were detectable. Consequently, the S. Enteritidis strains could only express fliC gene resulting in phase H1 flagellin. The examined S. Gallinarum strains were also demonstrated to have a cryptic flagellin gene (fliC). On the other hand, PCR results on S. Abortusequi (n = 2) indicated that both flagellin genes (fliC, fljB) and the whole phase variation system were present in both strains tested but only the H2 phase gene (fljB) was expressed. The phenotype of these strains could be clarified by motility test and/or by classical flagellar serology. The findings are also substantiated by the results of serovar-specific PCR for S. Typhimurium and S. Enteritidis. In conclusion, the PCR system developed in this study proved to be suitable for characterisation of Salmonella flagellin genes and confirmed serological results regarding all S. Typhimurium, S. Hadar and S. Enteritidis strains. This system could also identify cryptic flagellar genes of S. Abortusequi and S. Gallinarum.

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Abortion in dairy cattle causes considerable economic losses to the dairy industry. Aborted fetuses and samples from the corresponding aborting dams from 12 dairy herds in Beijing were tested for 9 abortifacient infectious pathogens by PCR between 2008 and 2010. From a total of 80 abortion cases collected during this period, infectious agents were detected in 45 (56.3%) cases, 22 (48.9%) of which represented co-infections with two or three infectious agents. The detected pathogens included infectious bovine rhinotracheitis virus (36.3%) and Neospora caninum (31.3%), followed by bovine viral diarrhoea virus (7.5%), Brucella abortus (6.3%), Tritrichomonas foetus (5%) and Toxoplasma gondii (1.3%). Campylobacter fetus, Coxiella burnetii and Chlamydophila psittaci were not detected in any abortion case. Findings from this study indicated that infectious bovine rhinotracheitis virus and Neospora caninum were the main potential causes of abortions in Beijing dairy herds, whereas the bacterial pathogens were not, in contrast to reports from other countries. This is the first study to test nine abortifacient infectious agents by PCR at the same time, and it is also the first time to report the involvement of a variety of infectious agents in bovine abortion cases in China.

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Enniatins (ENs), produced by Fusarium species are a group of mycotoxins with antimicrobial, insecticidal (GROVE & POPLE, 1980) and phytotoxic activities. PCR based assays were applied for detecting enniatin-producing strains of Fusarium avenaceum, F. poae and F. sporotrichioides isolated from wheat seeds originated of 30 geographic localities of Hungary. All F. sporotrichoides strains and except two of all F. poae strains gave positive signal to esysp1 and esysp2 primers as well as all F. avenaceum isolates were positive to esya1 and esya2 primers indicating the ability to produce ENs. This is a first report of the enniatin producing ability of Fusarium species associated to wheat in Hungary.

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Campylobacter jejuni and C. coli are among the most important causes of acute diarrhoea in humans throughout the world. Poultry meat is a major source of Campylobacter infections. Sensitive detection methods are necessary to identify contaminated samples. Detection of campylobacters by culturing is slow and tedious, whereas PCR technology offers the potential for rapid and sensitive detection, however, it may be inhibited when used directly for food or pre-enriched food samples. Different methods for sample and/or DNA preparation were studied to find an optimal combination for sensitive PCR detection of C. coli in enrichment broth. Buoyant density centrifugation (BDC) prior to cell lysis improved PCR detection of C. coli by 100-1000-fold. Preston enrichment broth spiked with 101-102 CFU ml-1 was detected positive after 18 h of enrichment. Specific flaA PCR detection of C. coli in enrichment broth following BDC and simple heat lysis of the cells can be conducted within two working days. This study is a part of the undergoing development of a rapid and sensitive molecular procedure for specific detection of C. coli in foods.

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Adenoviral nucleic acid was detected by polymerase chain reaction (PCR) in formalin-fixed paraffin-embedded tissue samples of a cat that had suffered from disseminated adenovirus infection. The identity of the amplified products from the hexon and DNA-dependent DNA polymerase genes was confirmed by DNA sequencing. The sequences were clearly distinguishable from corresponding hexon and polymerase sequences of other mastadenoviruses, including human adenoviruses. These results suggest the possible existence of a distinct feline adenovirus.

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