Development of a sensitive detection system for Cryptosporidium in environmental samples

The identification of Cryptosporidium species and genotypes is necessary to determine sources of infection in outbreaks and the risk factors associated with their transmission. Few studies have applied isolation methods to field samples because of difficulties with detection of oocysts in environmental samples, particularly in soil and manure. The objective of this study was to develop an easy to use method which can be applied to field samples to rapidly detect the presence of Cryptosporidium parasites and identify their species. The assay included an oocyst recovery method combined with spin column DNA extraction, followed by PCR-hybridization for detection and a real-time PCR-melting curve analysis for species assignment. An internal positive control (IPC) was developed to determine the presence of PCR inhibitory substances. Two oocyst recovery methods, sodium chloride and sucrose flotation techniques were compared. Two commercial DNA extraction kits were performed using feces, soil and water samples each inoculated with different concentration of Cryptosporidium oocysts. Subsequently, methods were used to test field samples. The sucrose flotation method provided the greatest analytical sensitivity detecting as few as 10 oocysts. The PCR-hybridization detection limit was 10 oocysts for feces and soil, and less than 10 oocysts for water samples. IPC was positive for all inoculated and field samples indicating 0% PCR inhibition. Cryptosporidium species DNA samples were detected with the real-time PCR and were differentiated by the melting curve analysis. The results of this study demonstrate the potential of the assay system for rapid detection of Cryptosporidium parasites in environmental samples.     

Food sovereignty, urban food access, and food activism: contemplating the connections through examples from Chicago

The idea of food sovereignty has its roots primarily in the response of small producers in developing countries to decreasing levels of control over land, production practices, and food access. While the concerns of urban Chicagoans struggling with low food access may seem far from these issues, the authors believe that the ideas associated with food sovereignty will lead to the construction of solutions to what is often called the ??food desert?? issue that serve and empower communities in ways that less democratic solutions do not. In Chicago and elsewhere, residents and activists often see and experience racial and economic inequalities through the variety of stores and other food access sites available in their community. The connections between food access, respect, and activism are first considered through a set of statements of Chicagoans living in food access poor areas. We will then discuss these connections through the work and philosophy of activists in Chicago centered in food sovereignty and food justice. Particular focus will be placed on Growing Power, an urban food production, distribution, and learning organization working primarily in Milwaukee and Chicago, and Healthy South Chicago, a community coalition focused on health issues in a working class area of the city.     

Biochemical characterisation of some non fermenting, non arginine hydrolysing mycoplasmas of ruminants

The pattern and kinetics of substrate oxidation by type and recent field strains of Mycoplasma agalactiae, Mycoplasma bovis, Mycoplasma bovigenitalium and Mycoplasma ovine/caprine serogroup 11 were investigated by measurement of oxygen uptake. Metabolism of a range of organic acids, sugars and alcohols was detected. All the test strains were unable to oxidise sugars, glycerol and the organic acids, fumarate, malate and alpha-ketoglutarate (1 mM). All strains oxidised organic acid l-lactate, 2-oxobutyrate and pyruvate and demonstrated the ability to oxidise alcohols, particularly isopropanol, which was oxidised at a high rate and high affinity (0.5 mol/mol isopropanol). Its oxidation was consistent with acetone formation, which may be of important in relation to pathogenicity. All strains oxidised similar substrates, however differences were observed between strains in terms of the relative rates and kinetic values for some substrates.     

Vegetative Compatibility and Mycotoxin Chemotypes among Fusarium graminearum (Gibberella zeae) Isolates from Wheat in Argentina

Gibberella zeae (anamorph Fusarium graminearum) is the main pathogen causing Fusarium head blight of wheat in Argentina. The objective of this study was to determine the vegetative compatibility groups (VCGs) and mycotoxin production (deoxynivalenol, nivalenol and 3-acetyl deoxynivalenol) by F. graminearum populations isolated from wheat in Argentina. VCGs were determined among 70 strains of F. graminearum isolated from three localities in Argentina, using nitrate non-utilizing (nit) mutants. Out of 367 nit mutants generated, 41% utilized both nitrite and hypoxanthine (nit1), 45% utilized hypoxanthine but not nitrite (nit3), 9% utilized nitrite but not hypoxanthine (NitM) and 5% utilized all the nitrogen sources (crn). The complementations were done by pairing the mutants on nitrate medium. Fifty-five different VCGs were identified and the overall VCG diversity (number of VCGs/number of isolates) averaged over the three locations was 0.78. Forty-eight strains were incompatible with all others, thus each of these strains constituted a unique VCG. Twenty-two strains were compatible with other isolates and were grouped in seven multimembers VCGs. Considering each population separately, the VCG diversity was 0.84, 0.81 and 1.0 for San Antonio de Areco, Alberti and Marcos Juarez, respectively. Toxin analysis revealed that of the 70 strains of F. graminearum tested, only 90% produced deoxynivalenol, 10% were able to produce deoxynivalenol and very low amounts of 3-acetyldeoxynivalenol. No isolate produced nivalenol. The results indicate a high degree of VCG diversity in the F. graminearum populations from wheat in Argentina. This diversity should be considered when screening wheat germplasm for Fusarium head blight resistance.     

ULTRASOUND-ASSISTED DIAGNOSIS OF RENAL DYSPLASIA IN A 3-MONTH-OLD QUARTER HORSE COLT

A 3-month-old foal was presented for correction of bilateral angular limb deformities. Azotemia was detected as an incidental finding. Small, misshapened, hyperechoic kidneys with decreased corticomedullary demarcation were noted with ultrasonography. Additionally, the internal renal architecture was abnormal in that the intrarenal vessels and distant collecting system were not clearly seen in either kidney. Ultrasound-guided renal biopsy was suggestive of congenital renal dysplasia, which was later confirmed at necropsy. Clinical, sonographic, and pathologic features of equine renal dysplasia are discussed.     

Serological study of contagious agalactia in herds of goats in the Canary Islands

An indirect ELISA, using local strains of Mycoplasma agalactiae and Mycoplasma mycoides subspecies mycoides large colony (MmmLC), was applied to evaluate the seroprevalence of M agalactiae and MmmLC in flocks of goats on each of the Canary Islands. In total 3890 samples of serum were collected from 204 flocks. The results indicated that the seroprevalence of both organisms is high on all the islands; average values of 55 per cent and 67 per cent were recorded, respectively, for M agalactiae and MmmLC.     

PCR based differentiation between Streptococcus dysgalactiae subsp. equisimilis strains isolated from humans and horses

Streptococcus dysgalactiae subsp. equisimilis (SDSE) can be severely pathogenic in humans and is increasingly isolated from horses with respiratory, reproductive or other diseases, although it is often considered a commensal bacterium. Here a PCR protocol is described for identifying SDSE recovered from humans. A multiplex PCR targeting the 16S rRNA and the streptokinase precursor gene has been optimized for differentiating between SDSE strains isolated from humans and those isolated from horses. Previously, the sequence of the streptokinase precursor gene of SDSE recovered from horses has been found in two human cases of pneumonia in Japan.