Evaluation of a multiplex polymerase chain reaction assay for simultaneous detection of Rhodococcus equi and the vapA gene

OBJECTIVE: To evaluate sensitivity and specificity of a multiplex polymerase chain reaction (PCR) assay for simultaneous detection of Rhodococcus equi and differentiation of strains that contain the virulence-associated gene (vapA) from strains that do not. SAMPLE POPULATION: 187 isolates of R equi from equine and nonequine tissue and environmental specimens and 27 isolates of bacterial species genetically or morphologically similar to R equi. PROCEDURE: The multiplex PCR assay included 3 gene targets: a universal 311-bp bacterial 16S ribosomal RNA amplicon (positive internal control), a 959-bp R equi-specific target in the cholesterol oxidase gene (choE), and a 564-bp amplicon of the vapA gene. Duplicate multiplex PCR assays for these targets and confirmatory singleplex PCR assays for vapA and choE were performed for each R equi isolate. An additional PCR assay was used to examine isolates for the vapB gene. RESULTS: Results of duplicate multiplex and singleplex PCR assays were correlated in all instances, revealing high specificity and reliability (reproducibility) of the vapA multiplex assay. Of the pulmonary isolates from horses with suspected R equi pneumonia, 97.4% (76/78) yielded positive results for vapA. Seven of 50 (14%) human isolates of R equi yielded positive results for vapA. Six human R equi isolates and 1 porcine isolate yielded positive results for vapB. No isolates with vapA and vapB genes were detected. CONCLUSIONS AND CLINICAL RELEVANCE: The multiplex PCR assay is a sensitive and specific method for simultaneous confirmation of species identity and detection of the vapA gene. The assay appeared to be a useful tool for microbiologic and epidemiologic diagnosis and research.     

Efficacy of a long-acting formulation of ceftiofur crystalline-free acid for the treatment of naturally occurring infectious bovine keratoconjunctivitis

OBJECTIVE: To evaluate the efficacy of ceftiofur crystalline-free acid (CCFA) administered into the posterior aspect of an ear for treatment of corneal ulceration associated with naturally occurring infectious bovine keratoconjunctivitis (IBK). ANIMALS: 78 beef calves located at Sierra Foothills Field Station (SFS) and 52 calves located at a commercial dairy (CD). All calves were from 3 to 9 months old. PROCEDURE: At each site, calves were randomly allocated to 1 of 2 treatment groups by use of a block design determined by corneal ulcer size. A single dose of CCFA (6.6 mg of ceftiofur equivalents/kg, s.c.) was administered into the posterior aspect of a pinna. A second group of calves received a single dose of vehicle (0.03 mL/kg, s.c.; controls). Corneal ulcers were photographed, and clinical signs were assessed in calves every 3 to 4 days for 21 days. RESULTS: A positive treatment effect was detected at SFS. Results at the CD were inconclusive because ulcer healing occurred rapidly in control and CCFA-treated calves. At SFS, treatment with CCFA resulted in shorter mean healing times, smaller corneal ulcer surface area measurements, amelioration of ocular discharge and photophobia, and a 50% increase in the percentage of calves healed by day 14. After adjustment for initial corneal ulcer size, treatment with CCFA resulted in a 4-fold increase in the odds of corneal ulcer healing by day 14, compared with controls. CONCLUSIONS AND CLINICAL RELEVANCE: A single dose of CCFA administered into the posterior aspect of a pinna had a positive treatment effect against naturally occurring IBK in calves with corneal ulcerations.     

Quantitative impact assessment of spray coverage and pest behavior on contact pesticide performance

BACKGROUND: Standard procedures for evaluation of pesticide performance do not take into account pest behavioral response or incomplete spray coverage. In this study, a series of laboratory experiments was conducted with two non‐systemic miticides (propargite and hexythiazox) applied to cotyledon cotton plants, which were subsequently infested with spider mites. The results of these laboratory experiments are discussed through a comprehensive pest population dynamics model. RESULTS: When cotton leaves were submerged in miticide solutions, both miticides provided effective control of spider mites. In a two‐choice test it was demonstrated that propargite was repellent to spider mites, but not hexythiazox. Finally, the spray coverage on cotton plants was varied, and, for both miticides, significantly positive relationships between spray coverage and spider mite mortality were shown. However, propargite required higher spray coverage (20%) than hexythiazox (10%) to control spider mites. A theoretical model showed that, without repellency, the pesticide performance is positively correlated with target pest mobility. If the pesticide is repellent, the probability of exposure decreases, especially for a less mobile pest. CONCLUSION: With an experimental and theoretical modeling approach, it was demonstrated how the combination of behavioral avoidance and low spray coverage can markedly reduce pesticide performance. Copyright © 2012 Society of Chemical Industry     

Innate lymphoid cells promote anatomical containment of lymphoid-resident commensal bacteria

The mammalian intestinal tract is colonized by trillions of beneficial commensal bacteria that are anatomically restricted to specific niches. However, the mechanisms that regulate anatomical containment remain unclear. Here, we show that interleukin-22 (IL-22)-producing innate lymphoid cells (ILCs) are present in intestinal tissues of healthy mammals. Depletion of ILCs resulted in peripheral dissemination of commensal bacteria and systemic inflammation, which was prevented by administration of IL-22. Disseminating bacteria were identified as Alcaligenes species originating from host lymphoid tissues. Alcaligenes was sufficient to promote systemic inflammation after ILC depletion in mice, and Alcaligenes-specific systemic immune responses were associated with Crohn's disease and progressive hepatitis C virus infection in patients. Collectively, these data indicate that ILCs regulate selective containment of lymphoid-resident bacteria to prevent systemic inflammation associated with chronic diseases.     

Clinical efficacy of the acyclic nucleoside phosphonate 9-(2-phosphonylmethoxypropyl)-2,6-diaminopurine (PMPDAP) in the treatment of feline immunodeficiency virus-infected cats

In in vitro studies, the acyclic nucleoside phosphonate 9-(2-phosphonylmethoxypropyl)-2,6-diaminopurine (PMPDAP) inhibited the replication of feline immunodeficiency virus (FIV). No information about its clinical efficacy is available so far. The aim of this prospective placebo-controlled, double-blinded study was to evaluate the antiviral efficacy of PMPDAP in cats naturally infected with FIV. Twenty cats were randomly assigned to two treatment groups receiving either PMPDAP (25 mg/kg) or placebo twice per week subcutaneously for 6 weeks. The general health status (Karnofsky's score), clinical signs, laboratory, immunological, and surrogate parameters were evaluated. No significant differences were found between PMPDAP- and placebo-treated cats, although cats treated with PMPDAP showed a tendency for improvement in their Karnofsky's score and clinical signs. Haematological side effects were noted in the PMPDAP-treated cats. Thus, PMPDAP may be an option in treating cats if it becomes available for veterinarians, but side effects have been monitored.     

The severity of smouldering peat fires and damage to the forest soil

Smouldering wildfires propagate slowly through surface and subsurface organic layers of the forest ground and severely affect the soil, producing physical, chemical and biological changes. These effects are caused by the prolonged heating and the large loss of soil mass but are poorly documented in the literature. A series of smouldering experiments with boreal peat have been conducted under laboratory conditions to quantify these effects using small-scale samples. Peat samples of 100 mm by 100 mm in cross section and 50 mm in depth of different moisture were exposed to an external ignition source. Thermocouples placed throughout the sample bed measured the temperature evolution and tracked the peat ignition, intensity and spread of the smouldering front. The results show that moisture content controls peat ignition and that moisture below 125 ± 10% (in dry base) are required. The severity of the smouldering peat on the soil has been quantified in terms of temperature vs. residence time curves and mass loss. The measurements show temperatures in excess of 300 °C for residence times of 1 h leading to sterilization of the soil and mass loss in burnt layers above 90%.     

Prevalence of Ehrlichia canis, Anaplasma platys, Babesia canis vogeli, Hepatozoon canis, Bartonella vinsonii berkhoffii, and Rickettsia spp. in dogs from Grenada

To identify the tick-borne pathogens in dogs from Grenada, we conducted a serologic survey for Ehrlichia canis in 2004 (104 dogs) and a comprehensive serologic and molecular survey for a variety of tick-borne pathogens in 2006 (73 dogs). In 2004 and 2006, 44 and 32 dogs (42.3% and 43.8%) were seropositive for E. canis, respectively. In 2006, several tick-borne pathogens were identified by serology and PCR. DNA of E. canis, Anaplasma platys, Babesia canis vogeli, Hepatozoon canis, and Bartonella sp. were identified in 18 (24.7%), 14 (19.2%), 5 (7%), 5 (7%), and 1 (1.4%) dogs, respectively. Six (8.2%) dogs were seropositive for Bartonella vinsonii subsp. berkhoffii. All dogs were seronegative and PCR-negative for Rickettsia spp. Coinfection with two or three pathogens was observed in eight dogs. Partial 16S rRNA E. canis and A. platys sequences were identical to sequences in GenBank. Partial 18S rRNA gene sequences from the Grenadian H. canis were identical to each other and had one possible mismatch (ambiguous base) from H. canis detected from Spain and Brazil. Grenadian B. c. vogeli sequences were identical to B. c. vogeli from Brazil and Japan. All of the detected pathogens are transmitted, or suspected to be transmitted, by Rhipicephalus sanguineus. Results of this study indicate that dogs from Grenada are infected with multiple tick-borne pathogens; therefore, tick-borne diseases should be included as differentials for dogs exhibiting thrombocytopenia, leukopenia, fever, or lethargy. One pathogen, E. canis, is also of potential public health significance.