Stomoxys calcitrans (Linnaeus, 1758), the stable fly, is a hematophagous pest that affects animal livestock worldwide by lowering productivity and mechanically transmitting pathogens. This study evaluated the effectiveness of the Nzi trap and a modified version (with and without bait) for capturing stable flies on two dairy farms in southwestern Uruguay, under conditions of natural infestation by these flies. The study used a 4×4 Latin square design with five replicates conducted independently at each dairy farm. Traps at each dairy were emptied daily to identify and count stable flies before being rotated and reset at the next planned location. Among all configurations, standard Nzi traps captured the highest number of S. calcitrans compared to the other trap evaluated, regardless of whether an attractant was used. This study represents the first evaluation in South America of Nzi traps for S. calcitrans used in combination with a commercial bait. Findings indicate that Nzi traps are a potentially effective tool for capturing S. calcitrans at dairy farms.

This study was conducted during the southern hemisphere summer (January and February 2023) at two dairy farms 3.1 km apart (Farm 1: -34.369338º S -57.879860º W, Farm 2: -34.345090º S -57.895231º W, Colonia, Uruguay, Supplementary Figure S1). The sampling period was based on previous work by Pérez (2021), which showed a seasonal peak between December and February. These dairies were small-scale farms, with 60 (Farm 1) and 80 (Farm 2) lactating Holstein cows.  In previous years, farm owners reported severe stable fly infestations, which led to decreased weight gain, lower milk yields, and skin lesions in their cattle.

Traps, bait, and experimental design

Two traps were evaluated: a typical Nzi trap (Mihok, 2002) and a locally assembled modified version (dataset D1 and Figure S2 in the supplementary material). Both were constructed with a local blue vinyl fabric; the modified trap had the same true surface area as the Nzi but incorporated three side entrances. Each trap was tested with or without a new commercial bait for S. calcitrans (Adios Moscas- Stomoxys calcitrans^TM, Products P651-Lure, Batch: 175.731, Chemtica International S.A, Costa Rica). It contains two ingredients in a plastic bag: 1) 1-octen-3-ol (formulated as a solid tablet-like matrix) and 2) a proprietary mixture of other chemicals in powder form (not yet disclosed). The bait was reconstituted in water at room temperature (300-350 mL/per container). The attractant package was affixed to the upper section of each trap.

The experimental design followed a 4×4 Latin square arrangement with five replicates conducted independently at each farm. Four different trap types were evaluated: the standard Nzi trap, a modified Nzi trap, and both versions equipped with bait. At each farm, four fixed sampling points were established.  Sampling points were located near the dairy, with sites separated by more than 50 meters. Traps were rotated daily among the four points in such a way that each trap occupied each position once during a 4-day cycle. This rotation scheme was repeated five times (i.e., five full Latin square replicates), ensuring that each trap was tested at each location a total of five times. The same protocol was implemented identically at both farms, allowing for replication and comparison under different environmental conditions. At the end of the experiment, a total of 40 collections were obtained for each trap type (20 at each farm), resulting in 160 trap collections overall (4 trap types × 40 collections per trap).

Species identification

Identification of stable flies was carried out using a stereomicroscope (Zeiss, model Stemi 508, Jena, Germany) according to Zumpt (1973). For each capture, we classified and quantified the specimens.

Characterization of the blue fabric

All traps were made from an opaque, shiny vinyl as a local alternative to phthalogen blue IF3GM cotton. A FORS (Fiber Optic Reflectance Spectroscopy) spectrum for this blue vinyl indicated that the main colorant was similar but not identical to the spectrum of copper phthalocyanine (CuPc) or Pigment Blue 15 (Poldi and Caglio, 2013), the exception being a lower peak in the blue at 450 nm. Spectra were taken with an Ocean Optics USB-4000 spectrophotometer using a deuterium-halogen light source (Florida, USA) and a Spectralon WS-1-SL white standard (Labsphere, USA). Full details are provided in supplementary material D2 and Figure S3.

Statistical analysis

All analyses were performed using Generalized Linear Mixed Models with a negative binomial distribution using Navure software (NAVURE, 2023).  To select the best model, data normality and AIC/BIC fits were compared. Trap performance was evaluated based on the number of flies captured, with trap type (Nzi and modNzi), bait (with or without), and their interaction included as fixed factors, while farm, trap position, day, and week were included as random effects.  Differences were considered statistically significant at p < 0.05.  To address the distance hypothesis, the exact distance (m) from the dairy was included as a fixed quantitative factor in this model. To assess the stability of capture rates over time, an interaction term between distance and date was tested using a likelihood ratio (LR) test.