J APPL POULT RES 2008. 17:93-100. doi:10.3382/japr.2007-00044
© 2008 Poultry Science Association
Tank Mixes: Consequences of Using Insecticide and Disinfectant Mixtures to Reduce Flies and Bacteria
D. W. Watson1,
C. K. Boohene,
S. S. Denning and
S. M. Stringham
Department of Entomology, Box 7626, North Carolina State University, Raleigh 27695
Correspondence: 1 Corresponding author: Wes_Watson{at}ncsu.edu
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SUMMARY
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The use of disinfectants and insecticides to control pathogens and the insects that may harbor avian pathogens has become routine for on-farm biosecurity programs. It is commonplace for producers to wash, disinfect, and apply insecticides to poultry houses between flock cycles. Occasionally, the urgency to repopulate the houses limits the time producers have to adequately perform preflock sanitization of the premise. The use of tank mixes (i.e., combining insecticides and disinfectants in a single application) saves time and labor. This study examined the consequences of using insecticide and disinfectant used separately or as a mixture for the control of the bacterium Salmonella Typhimurium and the house fly (Musca domestica). Aldehyde + formalin- and aldehyde + glutaraldehyde/quaternary ammonium (DC&R and Synergize)-based disinfectants were effective against Salmonella Typhimurium at the label rate. Disinfectant classes quaternary ammonium (Tryad), iodine (Dyne-O-Might), and peroxymonosulfate (Virkon S) were ineffective at label rates or in mixtures with insecticides. House fly mortality was 100% for tetrachlorvinphos/vapona (Ravap), Spinosad (Elector), and cyfluthrin (Tempo) and above 92% for permethrin (Martin’s Permethrin) and tetrachlorvinphos (Rabon) insecticides. Permethrin efficacy was compromised by the addition of disinfectants in all cases except the glutaraldehyde/quaternary ammonium blend. Elector efficacy was reduced when blended with iodine or peroxymonosulfate disinfectant classes. Tempo insecticidal activity was compromised when mixed with Tryad and Virkon S. Ravap and Rabon efficacy against house flies was unchanged in mixture.
Key Words: fly management • house fly • pathogen management • Salmonella Typhimurium • biosecurity
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DESCRIPTION OF PROBLEM
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Disease prevention is the fundamental premise of on-farm biosecurity programs [1]. These programs have focused on new and re-emerging animal diseases: avian influenza, Newcastle disease, poult enteritis mortality syndrome [2, 3], but persistent infectious agents (e.g., Campylobacter jejuni, Listeria monocytogenes, and several Salmonella species) continue to impinge on the well-being of the poultry flock and consumers of poultry products [4, 5]. These agents are among the leading foodborne bacterial pathogens in the United States and worldwide [6]. For consumers, outbreaks of foodborne illness may be associated with many foods, yet contaminated meat, poultry, and egg products are the most frequently reported sources [6].
Many studies have demonstrated the need for and benefits of implemented biosecurity measures to reduce the prevalence and transmission of diseases and enteric pathogens on the farm and to the consumer [7–9]. A variety of identified risk factors include age of buildings, age of flocks, mortality disposal, restricted access, protective clothing, water sanitizers, sanitation program, boot washes, and reduced exposure to potential reservoirs.
Most disinfectants suspended in clean water effectively controlled most viruses and bacteria within 10 min of exposure, yet efficacy decreased in the presence of suspended organic matter [10]. Viral and bacterial resistance and cross-resistance to commercial disinfectants continue to concern the industry [11]. Despite thorough cleaning and disinfection, Salmonella contamination remained on interior surfaces in caged layer houses [12].
Arthropods, especially the house fly (Musca domestica L.), contribute to the transmission and maintenance of bacteria associated with foodborne illnesses during the preharvest interval. Half of all flies captured near poultry houses containing chickens naturally infected with Campylobacter were positive for the bacterium [5, 13]. These highly mobile insects are capable of long flight, readily move within and between poultry houses, and may spread infectious agents [14].
North Carolina recently conducted a pesticide use survey of 1,459 poultry producers (38.7% of 3,761 total) [15]. Producers ranked the house fly as a common and difficult insect to control. Pyrethroid insecticides were used most often against this pest.
Although the economic benefits of biosecurity programs have been evaluated targeting disease prevention, their implementation may be expensive and is not universally accepted [7]. For North Carolina, the use of disinfectants to control disease was routine among 71.5% of the producers, whereas 61.1% of the respondents disinfected their poultry houses after each flock cycle [15]. However, to save on labor costs and application time, 76.5% of producers prepared mixtures of insecticides and disinfectants in 1 spray tank for insect and microbial control in poultry facilities. Mixing insecticides and disinfectants in 1 spray tank for insect and pathogen control in poultry facilities compromises the efficacy of 1 or both of the chemicals [16]. Changes in the regulation of insecticides and disinfectants since 1987 demonstrated the need to reevaluate the efficacies of commonly used insecticides and disinfectants for their intended purpose and as mixtures.
In this study, we used Salmonella Typhimurium and the house fly to model the possible effects of disinfectant and insecticide mixtures for their intended uses.
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MATERIALS AND METHODS
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Salmonella culturing
A bioassay procedure was designed to demonstrate the effect of insecticide and disinfectant mixtures on the bacterium Salmonella Typhimurium. Salmonella Typhimurium was originally isolated from a poultry house and maintained in pure culture on Luria-Bertani (LB) [17] nutrient agar slants. Cultures were streaked for isolation on LB agar and incubated for 24 h at 37°C. Colonies were transferred to LB broth tubes and incubated for 24 to 28 h. Standard plate count was performed to determine the total number of cells in the stock culture.
Bacteriocidal Bioassay
The disinfectants tested [18] were: Synergize, Tryad, DC&R, Virkon-S, and Dyne-O-Might (Table 1
). Disinfectants were purchased from a local agricultural supply store and prepared as directed by label instructions. Two disinfectant classes, phenolics or cresylic acids, were not included in this test. Disinfectant solutions were measured for pH after preparation.
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Table 1. Brand name, active ingredient, directions for preparation, and activity spectrum of disinfectants and insecticides tested1
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Selected insecticides [19] (Table 1
) were among the most commonly used by the North Carolina poultry industry, including pyrethroids, cyfluthrin (Tempo) and permethrin (Martin’s Permethrin), and organophosphates, tetrachlorvinphos (Rabon) and tetra-chlorvinphos plus vapona (Ravap) and Spinosad (Elector). These insecticides were purchased from a local agricultural supply store and prepared according to label instructions. Spinosad (Elector) was provided by the manufacturer for inclusion in this study. Insecticide solutions were measured for pH after preparation.
Mixtures of insecticides and disinfectant solutions were prepared as equivalents and adjusted for volume to prevent dilution of either active ingredient. Solutions and mixtures were measured for pH after preparation.
Approximately 1 x 108 cells from the stock Salmonella culture were transferred to 2 mL of sterile LB broth and mixed by vortex. Each test disinfectant (300 µL), insecticide, or mixture was added to the Salmonella-inoculated tube, and the culture was mixed by vortex. Treatments were replicated 3 times. Tubes were incubated for 10 min, and a 30-µL aliquot was spread-plated onto prepared LB agar plates and incubated 36 h at 37°C.
House Fly Culturing
The house fly colony used in this study was originally isolated from wild flies collected from a dairy and maintained in colony continuously since 1982 without insecticide pressure. Fly pupae were harvested from the larval medium by floatation. Air-dried fly pupae were transferred to 250-cc specimen cups and placed in a 30 x 30 cm screened cage provisioned with food and water. Five days posteclosion, adult flies from the larger colony were anesthetized with CO2 and sorted into groups of 30 flies each and placed in separate specimen cups with screen lids. This was done to facilitate the transfer of flies to treated surfaces in a seamless manner. Each cup was provisioned with water as the bioassay was prepared.
House fly bioassay procedures used in this study were similar to those of Geden et al. [16]. Exterior plywood sheets (1 cm thick) were purchased from a local home improvement store. Plywood sheets were cut into 32 square boards each measuring approximately 30 x 30 cm. Boards were allowed to weather outdoors for 8 wk to simulate natural weathering and dissipate volatiles associated with construction adhesives.
Insecticidal Bioassay
Because we were testing insecticides against susceptible house flies, we assumed that the lowest permissible application rate would be sufficient to determine activity. Insecticides, disinfectants, and mixtures were prepared fresh and applied to the boards with a 750-mL trigger sprayer until runoff. The treated boards were air-dried for 1 h before commencing with the bioassay.
House Fly Exposure to Treated Boards
A circular embroidery hoop 14 cm in diameter was fitted with a mesh screen to prevent fly escape (Figure 1). A 15-cm length of duct tape (48 mm wide) was applied to the surface of the board to provide an untreated surface for the flies to recover from the anesthesia. Thirty adult house flies (5 d old) were anesthetized with CO2 and then placed on the untreated duct tape. The screened embroidery hoop was placed over the recovering flies, and 2 rubber bands were stretched from opposing corners of the board to hold the hoop in place. After flies had recovered, test boards were suspended vertically from pins inserted into a wooden framework. The location of each board on the framework was randomized, and the experiment was replicated 3 times. House fly mortality was recorded at 2 and 6 h postexposure. Fly mortality was calculated as a percentage of the total number of flies dying from exposure to the treatments. Percentage of mortality data were transformed for homogeneity using log (n + 1). Analysis of variance was used to compare house fly mortality between treatments. Data were analyzed using a 1-way ANOVA [20].
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Figure 1. House fly bioassay on treated plywood boards. Flies were anesthetized with CO2 and placed immediately on the untreated tape surface to recover. Fly mortality after exposure to treated surfaces (left) and untreated surfaces (right) were compared 2 and 6 h postexposure.
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RESULTS
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When the aldehyde/quaternary ammonium/formalin disinfectant (DC&R) was used as directed, it prevented the growth of Salmonella Typhimurium (Table 2). The DC&R remained active against Salmonella Typhimurium when mixed with Rabon, Ravap, Tempo, Permethrin, and Elector. Ravap did not lose insecticidal activity when mixed with DC&R (Table 3). Insecticide efficacy was compromised for Permethrin when mixed with DC&R.
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Table 2. Growth of Salmonella Typhimurium in nutrient broth after exposure to mixtures of disinfectants and insecticides or disinfectants alone1
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Table 3. Percentage of mortality (6 h postexposure) of house flies exposed to insecticides and mixtures of insecticides and disinfectants
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The iodine-based disinfectant Dyne-O-Might did not prevent the growth of Salmonella Typhimurium (Table 2). The addition of Dyne-O-Might reduced the insecticidal activity of Permethrin and Elector (Table 3). The disinfectant had no negative effects on Ravap, Rabon, and Tempo activity against house flies.
Synergize, an aldehyde/glutaraldehyde/quaternary ammonium blend disinfectant, prevented the growth of Salmonella Typhimurium when used according to label rates; however, it lost bactericidal activity when mixed with Ravap, Rabon, Tempo, Permethrin, and Elector insecticides (Table 2). Ravap, Permethrin, Rabon, Tempo, and Elector retained insecticidal activity against house flies when mixed with Synergize (Table 3).
The aldehyde/quaternary ammonium blend disinfectant, Tryad, did not prevent the growth of the test isolate of Salmonella Typhimurium, alone or in mixture with insecticides (Table 2). Insecticidal activity of Ravap, Rabon, and Elector were unaffected or improved when mixed with Tryad (Table 3). The efficacy of test pyrethroids, Tempo and Permethrin, was reduced as a consequence of mixing with Tryad.
Virkon S, a peroxymonosulfate disinfectant known for activity against viruses and bacteria, was not active against Salmonella Typhimurium alone or in insecticide mixes (Table 2). Insecticidal activity was unchanged for Rabon and Ravap (Table 3). Insecticidal activity of both pyrethroids and Elector was reduced when mixed with Virkon S.
All disinfectants tested were acidic or slightly acidic when prepared as directed except Tryad, with a pH of 11.45 (Table 4). All insecticides except Ravap were relatively neutral pH when prepared as directed (Table 4). Dyne-O-Might and Virkon S were acidic or slightly acidic in all preparations. Ravap was acidic (pH 2.50), and the addition of the insecticide lowered pH in all disinfectants except Dyne-O-Might. Rabon was slightly alkaline pH, and mixtures increased pH in most cases. Tempo, Permethrin, and Elector became acidic in most preparations except the alkaline disinfectant Tryad.
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DISCUSSION
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Three factors influence the efficiency of disinfectants—exposure time, surface characteristics, and organic matter. The harsh environment of a commercial poultry house is particularly hostile to disinfectants, owing to large expanses of porous surface and high organic matter. In our study, we attempted to simulate that difficult environment without confounding our results with the unpredictable variables of surface characteristics and high organic matter. To do so, we manipulated time of exposure and created a test environment more favorable to the bacterium. This was accomplished by suspending the Salmonella Typhimurium test culture in nutrient broth, a favorable environment for the growth of the bacterium, to provide optimal conditions for the bacterium to withstand the bactericidal effects of the disinfectant. In addition, the selected disinfectants were challenged with a short exposure time in that medium. Under these conditions, DC&R and Synergize prevented the growth of Salmonella Typhimurium after a 10-min exposure. After the disinfectant was mixed with an insecticide, the components making up the formulation (active and inert ingredients) resulted in the loss of bactericidal activity for Synergize. Interestingly, a previous study reported that DC&R was the disinfectant most frequently compromised by mixing with insecticides with the notable exception of Ravap [16]. Salmonella Typhimurium grew on culture medium with a neutral pH after being exposed to the mixtures. It is therefore likely that changes in pH did not contribute to the loss of bactericidal activity [16]. We postulate that extending the length of exposure would have increased bactericidal activity certainly for vegetative cells but also resilient bacteria. For example, quaternary ammonium and peroxide disinfectants were most effective against the gram-positive bacterium, Staphylococcus aureus, after a 20-min exposure [21]. Furthermore, the stage of development for the bacterium may influence the efficacy of the disinfectant. For example, spore-forming bacteria tend to be susceptible in the vegetative stage, but resistant spores formed by the bacterium late in the growth phase are more difficult to kill with a disinfectant [22].
The 2 insecticidal classes tested in our study are commonly used in poultry houses: organophosphates (tetrachlorvinphos/vapona and tetrachlorvinphos) and pyrethroids (cyfluthrin and permethrin). Elector (Spinosad) is a relatively new insecticide introduced to the poultry industry in 2005. Of the 2 organophosphates tested, Ravap (tetrachlorvinphos/vapona) was 100% effective against the susceptible strain of house flies. Ravap retained its insecticidal activity after being mixed with DC&R, Synergize, Tryad, and Virkon S. In contrast, Rabon (tetrachlorvinphos) was 94% effective for the control of house flies. Interestingly, Rabon insecticidal activity increased when mixed with the disinfectants. Enhanced insecticidal activity was noted for Rabon mixed with quaternary ammonium and formalin in the previous study [16]. For the pyrethroid insecticides, Tempo (cyfluthrin) was most efficacious, providing 100% fly mortality. There was no loss of efficacy when mixed with Dyne-O-Might but a notable loss of activity when mixed with Tryad or Virkon S. Approximately 92% of the house flies were killed from exposure to the pyrethroid insecticide Permethrin (permethrin). Permethrin insecticidal activity was compromised when mixed with DC&R, Dyne-O-Might, Tryad, and Virkon S but not Synergize. Elector was 100% effective against house flies in our tests when used alone. There was little or no loss of activity against flies when mixed with DC&R, Synergize, or Tryad. Insecticidal activity was compromised for mixtures of Elector and Dyne-O-Might or Virkon S. Change in pH likely contributed to the loss of insecticidal activity in susceptible house flies (Table 4). Porosity of wood surface may have also influenced insecticidal activity [16].
Realistically, house fly insecticide resistance profiles vary regionally and in some cases from farm to farm depending on insecticide use history. House flies collected from New York caged layer houses were resistant to tetrachlorvinphos (Rabon), with more than 40% of the exposed flies surviving [23]. Similarly, more than 50% of the house flies survived exposure to permethrin, whereas 35% of the flies survived cyfluthrin [23]. In light of these findings, producers should approach the use of disinfectant and insecticide mixtures with caution, understanding the practice may cause a lapse in biosecurity.
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CONCLUSIONS AND APPLICATIONS
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- Disinfectants DC&R and Synergize effectively killed the Salmonella Typhimurium field isolate at the label rate.
- Mixtures of DC&R and insecticides did not lose efficacy against Salmonella Typhimurium.
- House fly control was compromised when Permethrin, Tempo, and Elector insecticides were mixed with certain disinfectants.
- Insecticides and disinfectants should be used as directed by the label.
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ACKNOWLEDGMENTS
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This project was funded through USDA Cooperative State Research, Education, and Extension Service competitive grant no. NCV-VMCG-0016. We thank Elanco Animal Health (Greenfield, IN) for providing Spinosad for use in this study.
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REFERENCES AND NOTES
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- Insecticides: Tempo (Bayer Healthcare LLC, Shawnee Mission, KS), Martin’s Permethrin (Control Solutions Inc., Pasodena, TX), Rabon (Boehringer Ingelheim Vetmedica Inc., Elwood, KS); Ravap (Boehringer Ingelheim Vetmedica Inc.), Elector (Elanco Animal Health, Indianapolis, IN).
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SUMMARY
DESCRIPTION OF PROBLEM
