Mycobacterium bovis Infection in Holstein Friesian Cattle, Iran

To identify strains of Mycobacterium bovis circulating in Iran, we used region of difference, spoligotypes, and variable number tandem repeats to genotype 132 M. bovis isolates from Holstein Friesian cattle. Despite wide geographic origins, the strains were genetically homogeneous. Increased distribution of cattle herds and inadequate control measures may have contributed to strain dispersion

Comparative evaluation of LAMP and Nested-PCR for the diagnosis of bovine paratuberculosis

AbstractIntroductionMycobacterium avium subsp. paratuberculosis causes paratuberculosis (Johne’s disease), a systemic infection and chronic inflammation of the intestine that affects many species, including bovine. Infection is widespread in livestock, and human populations are exposed. A possible association between MAP infection and Crohn’s disease in humans has been also described. Effective control of paratuberculosis has hampered due to lake of rapid and accurate diagnostic test. Range of diagnostic tests is available, but all have inborn limitations. The present study was designed to develop a loop-mediated isothermal amplification (LAMP) assay for the rapid and simple detection of Mycobacterium avium subsp. paratuberculosis (MAP).Materials and methodsSix primers were specially designed for recognizing eight distinct sequence of insertion sequence 900 (IS900). To determine the sensitivity of the LAMP assay, 10-fold serial dilutions were made from 431ng/μl MAP stock solution and compared with Nested-PCR results obtained using similar templates at identical concentrations. Detection limit of the LAMP was defined as the last positive dilution and the reactions were performed four times to examine the reproducibility of the test. The specificity of the assays were evaluated by testing three Gram-positive bacteria including Mycobacterium bovis AN5, Mycobacterium tuberculosis DT and Mycobacterium avium avium.ResultsSensitivity of this assay for detection of DNA of MAP was 4fg/μl and the specificity was 100%. This assay successfully detected MAP not only in the bacterial cultures but also in clinical fecal samples and the specificity of both PCR was 100%. This LAMP method is performed under isothermal conditions and no special apparatus is needed. In addition, its reactivity is directly observed with the naked eye without electrophoresis either as turbidity or in the form of a color change when SYBR Green 1, a fluorescent dsDNA intercalating dye, is employed.ConclusionsThis assay is rapid which requires nearly 1h for detection of MAP, low in cost and simple to perform, sensitive and practical tool for the detection of MAP and will be useful in facilitating the early diagnosis of paratuberculosis (Johne’s disease) caused by the organism

Bovine tuberculosis in Iran: The past, present and future of a national disease control program

Bovine tuberculosis (bTB), caused by Mycobacterium bovis, is an important zoonotic disease with global distribution. While M. bovis is inherently resistant to pyrazinamide, human cases of infection with M. bovis might experience serious cure failures if no correct identification of the pathogen is achieved. Bovine TB was initially reported in Iran by a French veterinarian in local breeds of cattle. An official attempt to control the disease was started in the 1940s, which runs today on a national scale. This mini-review addresses a variety of different epidemiological issues in bTb control in the world and in Iran from an immunologist's eye to find the cure for human cases infected with M. bovis. In addition, the benefits and drawbacks of this control scheme are discussed

Genotyping analysis of bovine, ovine, and caprine paratuberculosis in Iran: An IS900-RFLP study

Objective/background: Mycobacterium avium subspecies paratuberculosis (MAP) has been reported in Iranian cattle since 1965. Little is known on the population genetics of MAP in this Middle-Eastern State. A principle scope of this study was therefore genetic characterization of MAP isolates collected from farm animals in Iran. Methods: Sixteen field isolates of MAP collected from bovine (n = 8), ovine (n = 3), and caprine (n = 2) hosts in Alborz, Fars, Isfahan, Qazvin, Tehran, and Zanjan provinces were subcultured on mycobactin J-supplemented Herrold's egg slopes in order to provide the required genomic material. The laboratory strain MAP III & V was included as the reference strain. IS900-RFLP (Restriction Fragment length Polymorphism) was conducted using BsteII restriction enzyme. Application of IS900-RFLP genotyping on 16 Iranian MAP isolates in the present study classified them into six observable but similar types represented by two clustered and four orphan types. The laboratory strain MAP III & V displayed a totally different pattern easily distinguishable from that of Iranians. Results: Detection of six genotypes among 16 wild isolates is an indication of a population with a potentially high level of diversity. We assume that, with inclusion of more field isolates, it is very likely that even higher diversity may be observed within the studied isolates. The different patterns displayed by the Iranians and the laboratory strain in this work might explain the independent evolutionary pathways these have gone through to evolve from their ancestral clones. Conclusion: Further description on population genetic of MAP in Iran urges for more epidemiological work using similar and alternative standard genotyping system

Mycobacterial coinfection and persisting bovine tuberculosis—Has the time arrived for a policy review?

Objective/Background: Bovine tuberculosis (BTb) is mainly a disease of cattle, although it continues to infect human populations across the world. Operation of a test and slaughter plan in Iran since 1981 has lowered the frequency of BTb from >5% to <0.14% at the national scale. In 2015, unusual uncontrollable epidemics of BTb were detected in two cattle farms in municipal suburbs of Qazvin and Isfahan. These farms had a tuberculin-test-certified record of BTb-free status for the past 5 consecutive years, with no new cattle registered with either of the two herds during this time period. Routine tuberculination of the bovids in 2015 resulted in the detection of tuberculin-positive animals that were subsequently removed from the herds. Serial tuberculin tests improved the situation, as new reactors were found each time. The aim of this research is based on isolation and identification of Mycobacterium from infected animals in both farms. Methods: To investigate the situation, major mesenteric/mediastinal lymph nodes from the culled reactor animals along with specimens from bulk milk tanks, trapped rats living on the farms, and environmental specimens were collected and subjected to bacterial culture. Tuberculin-positive cattle were also subjected to paratuberculosis enzyme-linked immunosorbent assay (ELISA), ESAT-6 ELISA, and gamma-interferon tests. Results: In bacterial culture, Mycobacterium bovis, Mycobacterium microti, and Mycobacterium avium subsp. paratuberculosis were isolated from collected specimens at both farms. Conclusion: There is circumstantial evidence supported by previous studies to expect a high frequency of M. avium subsp. paratuberculosis infection in Iranian cattle/sheep farms. This observation might explain the large skin reaction size seen at the avian tuberculin injection site in tested animals in these farms. Introduction of a third infection with M. microti, possibly by rodents visiting the farms, might have triggered immunological reactions that have ended the surge of BTb. If correct, we assume that a technical review of the Iranian test and slaughter scheme against BTb is required to address persisting cases of BTb in disease-free farms, as described here

Frequency of mycobacterial infections in retailed fish in Karaj: An Iranian perspective

Objective/Background: Mycobacterium species comprise one of the most frequently-reported causative agents in granulomatous lesions of fish. Also, mycobacteria have been documented as fish-born zoonoses. Ornamental aquaria, commercial aquaculture, and wild fisheries therefore produce a potential risk of infection for humans through physical contact and consumption of fishes. A number of fish mycobacteriosis and fish-born zoonotic cases have been reported to date in Iran. Methods: In order to examine the frequency of mycobacteria existence in fish supplied to the public in seafood retail outlets, whole fresh fish from cold water (n = 50) and tropical (n = 50) audible fish were obtained from five stores in Karaj, the central city of Alborz province. The head, tail, and offal of these fish were sampled during the necropsy and used for mycobacterial culture on Lowenstein–Jensen slopes. Results: In total, 15 acid-fast isolates were collected including 10 from tropical fish. The 16S ribosomal RNA and hsp genes of all isolates were polymerase chain reaction amplified and sequenced. Mycobacterium fortuitum was the most frequent mycobacterium identified in the study panel according to the Basic Local Alignment Search Tool search results. Work is still ongoing to characterize the other cultured mycobacterial isolates. Conclusion: The detection of 15 culturable mycobacterial isolates from 100 audible fish is an indication of bacteria highly active in the colonization in fish populations. Assessing the potential zoonotic risk raised from exposure of human to fishes in Iran demands search for evidence of linkages between mycobacteria infecting human cases and fishes

Occurrence of bovine TB in Iranian cattle herds of Khuzestan, a laboratory study on postmortem specimens from tuberculin-positive cows

Khuzestan is a southwestern province of Iran with proximity to the Persian Gulf and the international border with Iraq where harsh climate seriously affects this oil-rich region. In a search for causative agents of bovine tuberculosis (bTB), slaughterhouse specimens (lymph nodes) from 32 tuberculin-positive cows originating from 17 farms were cultured on Lowenstein–Jensen slopes. This was further extended with bacterial culture of postmortem material from 6 trapped feral mice straying on the same farm premises. Twenty-five bovine and 2 murine acid-fast isolates were consequently obtained with all of them confirmed as Mycobacterium tuberculosis (MTB) complex bacteria by IS6110-PCR experiment. Spoligotyping and RD4 typing of the 2 murine and some of the collected bovine isolates left no doubt that Mycobacterium bovis is the principle and possibly the single culprit in bTB in this region. It does not come as a surprise as previous exhaustive works have shown in the Iranian environment other members of MTB complex than M. bovis are very unlikely to have any role in the epidemiology of bTB

Mixed infection zones may be important in the epidemiology of contagious agalactia

Introduction: The current study was designed to detect Mycoplasma agalactiae (Ma), Mycoplasma mycoides subsp. capri (Mmc), Mycoplasma capricolum subsp. capricolum (Mcc) and Mycoplasma putrefaciens (Mp) in sheep and goats with clinical signs consistent with contagious agalactia