Claudia Muñoz-Zanzi

Identification of a Sporozoite-Specific Antigen from Toxoplasma gondii

Abstract Reduction of risk for human and food animal infection with Toxoplasma gondii is hampered by the lack of epidemiological data documenting the predominant routes of infection (oocyst vs. tissue cyst consumption) in horizontally transmitted toxoplasmosis. Existing serological assays can determine previous exposure to the parasite, but not the route of infection. We have used difference gel electrophoresis, in combination with tandem mass spectroscopy and Western blot, to identify a sporozoite-specific protein (T. gondii embryogenesis-related protein [TgERP]), which elicited antibody and differentiated oocyst- versus tissue cyst–induced infection in pigs and mice. The recombinant protein was selected from a cDNA library constructed from T. gondii sporozoites; this protein was used in Western blots and probed with sera from T. gondii–infected humans. Serum antibody to TgERP was detected in humans within 6–8 mo of initial oocyst-acquired infection. Of 163 individuals in the acute stage of infection (anti–T. gondii IgM detected in sera, or <30 in the IgG avidity test), 103 (63.2%) had detectable antibodies that reacted with TgERP. Of 176 individuals with unknown infection route and in the chronic stage of infection (no anti–T. gondii IgM detected in sera, or >30 in the IgG avidity test), antibody to TgERP was detected in 31 (17.6%). None of the 132 uninfected individuals tested had detectable antibody to TgERP. These data suggest that TgERP may be useful in detecting exposure to sporozoites in early T. gondii infection and implicates oocysts as the agent of infection.

Pooled-Sample Testing as a Herd-Screening Tool for Detection of Bovine Viral Diarrhea Virus Persistently Infected Cattle

The study was conducted to develop methodology for least-cost strategies for using polymerase chain reaction (PCR)/probe testing of pooled blood samples to identify animals in a herd persistently infected with bovine viral diarrhea virus (BVDV). Cost was estimated for 5 protocols using Monte Carlo simulations for herd prevalences of BVDV persistent infection (BVDV-PI) ranging from 0.5% to 3%, assuming a cost for a PCR/probe test of

Toxoplasma gondii Oocyst-specific antibodies and source of infection

20. The protocol associated with the least cost per cow involved an initial testing of pools followed by repooling and testing of positive pools. For a herd prevalence of 1%, the least cost per cow was

Evaluation of the sensitivity of reverse-transcription polymerase chain reaction to detect porcine reproductive and respiratory syndrome virus on individual and pooled samples from boars.

2.64 (95% prediction interval =

Leptospira Contamination in Household and Environmental Water in Rural Communities in Southern Chile

1.72,

A novel method of analyzing daily milk production and electrical conductivity to predict disease onset.

3.68), where pool sizes for the initial and repooled testing were 20 and 5 blood samples per pool, respectively. Optimization of the least cost for pooled-sample testing depended on how well a presumed prevalence of BVDV-PI approximated the true prevalence of BVDV infection in the herd. As prevalence increased beyond 3%, the least cost increased, thereby diminishing the competitive benefit of pooled testing. The protocols presented for sample pooling have general application to screening or surveillance using a sensitive diagnostic test to detect very low prevalence diseases or pathogens in flocks or herds.

Household Characteristics Associated with Rodent Presence and Leptospira Infection in Rural and Urban Communities from Southern Chile

Infection source can determine cost-effective public health interventions. To quantify risk of acquiring Toxoplasma gondii from environmental sources versus from meat, we examined serum from pregnant women in Chile. Because 43% had oocyst-specific antibodies, we conclude that contaminated meat remains the primary source of infection but that environmental sources also pose substantial risk.

A method to provide improved dose–response estimates for airborne pathogens in animals: An example using porcine reproductive and respiratory syndrome virus

Boar studs are continuously monitored for the presence of porcine reproductive and respiratory syndrome virus (PRRSV) by testing different biological samples by reverse-transcription polymerase chain reaction (RT-PCR). In most cases, samples are run in pools, even though the impact of pooling on the sensitivity of RT-PCR is unknown. The objective of this study was to evaluate the feasibility of using PCR on pooled samples through the estimation of the sensitivity of RT-PCR on different biological samples run individually, in pools of 3 and in pools of 5. Twenty-nine boars were inoculated with a low virulent PRRSV isolate. Serum, blood swab, and semen samples were obtained from each boar every 2 to 3 days for 2 weeks. Each sample was tested by RT-PCR undiluted or diluted 1:3 and 1:5 with negative samples. Eleven of the 29 boars did not appear to get infected from the inoculum, as evidenced by no seroconversion 15 days after inoculation. Data from the other 18 boars showed that serum was the best sample to detect PRRSV during acute infection, with the blood swab sample performing almost as well. Semen samples failed to detect PRRSV infection in most of the cases. Pooling samples at pool sizes of 3 and 5 resulted in a decrease in the sensitivity of RT-PCR. Sensitivity was reduced by 6% and 8%, respectively, when serum or blood swab samples were run in pools of 5. The impact of pooling on the sensitivity of PCR was higher in samples taken during the beginning of the viremic period.

Toxoplasma gondii in feral American minks at the Maullín river, Chile.

Leptospirosis is a zoonosis of global distribution that affects tropical and temperate areas. Under suitable conditions, Leptospira can survive in water and soil and contribute to human and animal infections. The objective of this study was to describe the presence of pathogenic Leptospira in peri-domestic water samples from rural households in southern Chile. Water samples, including puddles, containers, animal troughs, rivers, canals, and drinking water were collected from 236 households and tested for Leptospira using a PCR assay targeting the lipL32 gene. Evidence of Leptospira presence was detected in all sample types; overall, 13.5% (77/570) samples tested positive. A total of 10/22 (45.5%) open containers, 12/83 (14.5%) animal drinking sources, 9/47 (19.1%) human drinking sources, and 36/306 (19.3%) puddles tested positive. Lower income (OR = 4.35, p = 0.003), increased temperature (OR = 1.23, p < 0.001), and presence of dogs (OR = 15.9, p = 0.022) were positively associated with positive puddles. Increased number of rodent signs was associated with positive puddles in the household (OR = 3.22); however, only in the lower income households. There was no association between PCR positive rodents and puddles at the household level. Results revealed the ubiquity of Leptospira in the household environment and highlight the need to develop formal approaches for systematic monitoring.

Seroepidemiology of leptospirosis in dogs from rural and slum communities of Los Rios Region, Chile

This study evaluates the changes in milk production (yield; MY) and milk electrical conductivity (MEC) before and after disease diagnosis and proposes a cow health monitoring scheme based on observing individual daily MY and MEC. All reproductive and health events were recorded on occurrence, and MY and MEC were collected at each milking from January 2004 through November 2006 for 587 cows. The first 24 mo (January 2004 until December 2005) were used to investigate the effects of disease on MY and MEC, model MY and MEC of healthy animals, and develop a health monitoring scheme to detect disease based on changes in a cows MY or MEC. The remaining 11 mo of data (January to November 2006) were used to compare the performance of the health monitoring schemes developed in this study to the disease detection system currently used on the farm. Mixed model was used to examine the effect of diseases on MY and MEC. Days in milk (DIM), DIM x DIM, and ambient temperature were entered as quantitative variables and number of calves, parity, calving difficulty, day relative to breeding, day of somatotropin treatment, and 25 health event categories were entered as categorical variables. Significant changes in MY and MEC were observed as early as 10 and 9 d before diagnosis. Greatest cumulative effect on MY over the 59-d evaluation period was estimated for miscellaneous digestive disorders (mainly diarrhea) and udder scald, at -304.42 and -304.17 kg, respectively. The greatest average daily effect was estimated for milk fever with a 10.36-kg decrease in MY and 8.3% increase in MEC. Milk yield and MEC was modeled by an autoregressive model using a subset of healthy cow records. Six different self-starting cumulative sum and Shewhart charting schemes were designed using 3 different specificities (98, 99, and 99.5%) and based on MY alone or MY and MEC. Monitoring schemes developed in this study issue alerts earlier relative to the day of diagnosis of udder, reproductive, or metabolic problems, are more sensitive, and give fewer false-positive alerts than the disease detection system currently used on the farm.