Serological and molecular diagnosis of paratuberculosis in dairy cattle

In this study, the presence of paratuberculosis was investigated by serological and molecular methods in herds of dairy cattle. Blood, milk, and stool samples of 147 cows aged 2 years old or older with chronic diarrhea were collected. A California mastitis test (CMT) was performed on milk samples. Indirect paratuberculosis enzyme-linked immunosorbent assay (ELISA) test was used for serological investigation. Polymerase chain reaction (PCR) was utilized for molecular identification of Mycobacterium avium subsp. paratuberculosis (MAP) from milk and stool samples. Immunomagnetic separation (IMS) method was used for milk and fecal samples. Eighteen (12.24%), 44 (29.93%), 36 (24.49%), 28 (19.05%), and 21 (14.29%) of 147 milk samples were negative, suspicious, CMT (+), CMT (++), and CMT (+++) by CMT, respectively. According to ELISA results, 18 (12.24%) serum samples were positive. In PCR of milk and stool samples, MAP DNA was detected in 20 (13.61%) and 42 (28.57%) of samples, respectively. In IMS PCR assays of the same samples, positivity was not detected. In this study, paratuberculosis was found at high rates in Kayseri. In conclusion, it was detected that mastitis symptoms in paratuberculosis were subclinical and not always observed, and the use of diagnostic laboratory methods may be an important aid in revealing diseases

Prevalence and distribution of Arcobacter species in various sources in Turkey and molecular analysis of isolated strains by ERIC-PCR

Aims: To determine the prevalence of Arcobacter in various food, animal and water sources in Turkey and to subtype the isolated strains using enterobacterial repetitive intergenic consensus (ERIC)-PCR.Aims: To determine the prevalence of Arcobacter in various food, animal and water sources in Turkey and to subtype the isolated strains using enterobacterial repetitive intergenic consensus (ERIC)-PCR.Methods and Results: A total of 806 samples consisting of chicken (100) and turkey meat (100); minced beef (27); rectal swabs from cattle (173), sheep (68) and dogs (62); cloacal swabs of broilers (100) and layers (100); gall bladders of cattle (50) and drinking water samples (26) were examined. A previously described membrane filtration method was used for the isolation. Isolates were identified at species level using multiplex-PCR and discriminated by ERIC-PCR for subtyping. Ninety-eight (12·1%) of the samples examined were found positive for arcobacters. Arcobacter spp. were isolated from 68%, 4%, 6·9%, 8% and 37% of chicken and turkey meats, rectal swabs and gall bladders of cattle and minced beef, respectively. No arcobacters were obtained from the rectal swabs of sheep and dogs, cloacal swabs of broilers and layers, and water samples examined. In total, 99 Arcobacter isolates were obtained. Of these isolates, 92 were identified as Arcobacter butzleri, five were Arcobacter skirrowii and two were Arcobacter cryaerophilus. Thirteen distinct DNA profiles among A. butzleri isolates were obtained by the ERIC-PCR. Of these profiles, eight were from chicken carcass, three from cattle rectal swab and two from minced beef meat isolates. Some of the isolates originated from different sources gave the same DNA profiles. All isolates of A. skirrowii and A. cryaerophilus gave different DNA profiles.Conclusions: Poultry carcasses, minced beef meat, rectal swabs and gall bladders of cattle were found to be positive for Arcobacter spp. A. butzleri was the predominant species isolated. In addition, large heterogeneity among the Arcobacter isolates was determined.Significance and Impact of the Study: Contamination of the poultry carcasses and minced beef meat, rectal and gall bladder samples of cattle with arcobacters poses a risk for both human and animal infections. Detection of several different Arcobacter strains may suggest multiple sources for contamination and infection

Molecular typing and cdt genes prevalence of Campylobacter jejuni isolates from various sources

The genetic diversity of 168 Campylobacter jejuni isolates originating from human (n=30), cattle (n=36), sheep (n=44), dog (n=35), and poultry (n=21) and cdt genes prevalence of the isolates were investigated. To determine the genetic diversity of these strains, random amplified polymorphic DNA-polymerase chain reaction (PCR) using a random primer (M13) was performed. The numbers of genotypes determined in human, cattle, sheep, dog, and poultry isolates were 19, 18, 17, 18, and 6, respectively. To find out the prevalence of cdt genes in C. jejuni isolates simultaneously, a multiplex PCR was performed. The prevalence of the separate cdt genes was found to vary from 69% to 100% for cdtA, 92% to 100% for cdtB, and 39% to 98% for cdtC. These rates without host discriminating were 95%, 98%, and 93% for cdtA, cdtB, and cdtC, respectively. The prevalence of all three cdt genes in strains originating from human, cattle, sheep, dog, and poultry were 87%, 67%, 84%, 89%, and 39%, respectively. These results showed the relatively high genetic heterogeneity and variation of cdt genes among C. jejuni isolates from various sources except for poultry isolates. This study gives baseline data on molecular characterization of C. jejuni strains from different sources

The prevalence of Campylobacter jejuni in various sources in Kayseri, Turkey, and molecular analysis of isolated strains by PCR-RFLP

The objective of this study was to isolate, identify, and genotype Campylobacter jejuni from various sources in the province of Kayseri, Turkey. A total of 6667 samples consisting of 5167 human fecal swabs, 600 dog rectal swabs, 600 cattle gallbladders, and 300 chicken carcasses were examined. The samples were plated onto mCCDA (cefoperazone charcoal desoxycholate agar) agar. In order to identify C. jejuni, phenotypic tests and PCR (polymerase chain reaction) were performed. C. jejuni was isolated in 1.43%, 43.50%, 31.16%, and 56% of the human, dog, cattle, and chicken samples, respectively. Among the 690 C. jejuni strains that were isolated during the study period, 200 C. jejuni strains (50 strains from each species) were randomly selected. The selected strains were typed by using PCR-RFLP (polymerase chain reaction-restriction fragment length polymorphism) fla-typing. DdeI and HinfI restriction enzymes were used for molecular typing. Following the DdeI enzyme application, the strains produced various numbers of bands between 4 and 7, with a total of 20 different band profiles. No similar band profiles were seen among the strains isolated from different sources. It was found that HinfI was not a more discriminative enzyme for fla-typing of C. jejuni isolates

Characterization of Mono- and Mixed-Culture Campylobacter jejuni Biofilms

Campylobacter jejuni , one of the most common causes of human gastroenteritis, is a thermophilic and microaerophilic bacterium. These characteristics make it a fastidious organism, which limits its ability to survive outside animal hosts. Nevertheless, C. jejuni can be transmitted to both humans and animals via environmental pathways, especially through contaminated water. Biofilms may play a crucial role in the survival of the bacterium under unfavorable environmental conditions. The goal of this study was to investigate survival strategies of C. jejuni in mono- and mixed-culture biofilms. We grew monoculture biofilms of C. jejuni and mixed-culture biofilms of C. jejuni with Pseudomonas aeruginosa . We found that mono- and mixed-culture biofilms had significantly different structures and activities. Monoculture C. jejuni biofilms did not consume a measurable quantity of oxygen. Using a confocal laser scanning microscope (CLSM), we found that cells from monoculture biofilms were alive according to live/dead staining but that these cells were not culturable. In contrast, in mixed-culture biofilms, C. jejuni remained in a culturable physiological state. Monoculture C. jejuni biofilms could persist under lower flow rates (0.75 ml/min) but were unable to persist at higher flow rates (1 to 2.5 ml/min). In sharp contrast, mixed-culture biofilms were more robust and were unaffected by higher flow rates (2.5 ml/min). Our results indicate that biofilms provide an environmental refuge that is conducive to the survival of C. jejuni

Fe(III) Reduction and U(VI) Immobilization by Paenibacillus sp. Strain 300A, Isolated from Hanford 300A Subsurface Sediments

A facultative iron-reducing [Fe(III)-reducing] Paenibacillus sp. strain was isolated from Hanford 300A subsurface sediment biofilms that was capable of reducing soluble Fe(III) complexes [Fe(III)-nitrilotriacetic acid and Fe(III)-citrate] but unable to reduce poorly crystalline ferrihydrite (Fh). However, Paenibacillus sp. 300A was capable of reducing Fh in the presence of low concentrations (2 μM) of either of the electron transfer mediators (ETMs) flavin mononucleotide (FMN) or anthraquinone-2,6-disulfonate (AQDS). Maximum initial Fh reduction rates were observed at catalytic concentrations (<10 μM) of either FMN or AQDS. Higher FMN concentrations inhibited Fh reduction, while increased AQDS concentrations did not. We also found that Paenibacillus sp. 300A could reduce Fh in the presence of natural ETMs from Hanford 300A subsurface sediments. In the absence of ETMs, Paenibacillus sp. 300A was capable of immobilizing U(VI) through both reduction and adsorption. The relative contributions of adsorption and microbial reduction to U(VI) removal from the aqueous phase were ∼7:3 in PIPES [piperazine- N , N ′ - bis(2-ethanesulfonic acid)] and ∼1:4 in bicarbonate buffer. Our study demonstrated that Paenibacillus sp. 300A catalyzes Fe(III) reduction and U(VI) immobilization and that these reactions benefit from externally added or naturally existing ETMs in 300A subsurface sediments

Seroprevalence of Bovine Leptospirosis in Kayseri, Turkey and Detection of Leptospires by Polymerase Chain Reaction

This study was carried out to determine the seroprevalence of bovine leptospirosis in Kayseri, Turkey and to detect the leptospires in blood and urine of clinically suspect animals by serologic and molecular methods. A total of 2395. blood samples were collected from slaughterhouses located during 12 months (from May 2005 to April 2006) determine the seroprevalence. In addition, blood and urine samples were collected from 500 clinically leptospirosis suspect cattle. Microscopic Agglutination Test (MAT) and Enzyme Linked Immunosorbent Assay (ELISA) were used as serological tests. Polymerase Chain Reaction (PCR) was used for molecular examinations. In the serological analysis of 2395 blood samples collected from slaughterhouses, 609 (25.42%) and 433 (18.07%) samples were found to be positive by the MAT and ELISA, respectively. Seven (1.40%) out of 500 leptospirosis suspect cattle were found to be infected by the MAT and ELISA and Leptospira interrogans serovar Hardjo and L. kirchneri serovar Grippotyphosa were the predominant serovars. And also leptospires were detected in the urine samples of these 7 cattle (1.40%) by PCR. No agent was detected in the blood of suspect animals by PCR. The results of this study have shown that leptospirosis is highly prevalent and predominantly caused by L. interrogans serovar Hardjo and L. kirchneri serovar Grippotyphosa in this region. Because PCR can compete to the serological tests for detecting the leptospires in urine samples of suspect animals, the molecular analysis may contribute to the diagnosis of the infection

Conventional and molecular biotyping of Brucella strains isolated from cattle, sheep and human

In this study, the role of Brucella spp. in cattle and sheep abortions among Kayseri region was investigated and predominant subspecies and biovars in this region were determined by conventional and molecular biotyping methods. For this purpose, 61 cattle and 64 sheep abortion material and also 50 human blood isolates were examined. A total of 29 Brucella spp. 17 (27.9%) and 12 (18.7%) of which were isolated from cattle and sheep specimens, respectively) were isolated from animal sources. Both animal and human isolates were typed by conventional and Enhanced AMOS-ERY PCR methods. All Brucella spp. strains isolated from cattle were found to belong to B. abortus biovar 3 and biovar 3b using conventional and molecular typing methods, respectively. All sheep originated Brucella spp. strains and human originated Brucella spp. strains were found to belong to B.melitensis biovar 3 using both conventional and molecular methods. As a result, predominant biovars causing brucellosis in human, cattle and sheep in Kayseri, Turkey were detected. These findings were considered to be useful in prevention and controlling activities for Brucellosis in Turkey