**Abstract**

American foulbrood (AFB) is the most severe bacterial disease that affects honey bees, having a nearly cosmopolitan distribution. AFB's causative agent is *Paenibacillus larvae*. AFB kills infected honey bee larvae; however, it eventually leads to the collapse of the entire colony when left untreated. The infection takes place by the ingestion of the spores with the food provided by adult worker bees to the larvae. In South America (SA) the disease was first described in 1989 in Argentina, constituting the first sanitary challenge for beekeepers to overcome. Prevention and control measures of AFB in SA countries generally include vigilance for early diagnosis, isolation of apiaries with cases of AFB, and multiplication of healthy colonies with hygienic queens, among others. The extensive use of tetracycline hydrochloride in Argentina has led to the development of resistant *P. larvae* isolates. In this context, the development of alternative and effective methods for the control and prevention of AFB disease is crucial. Currently, alternative strategies for the prevention and treatment of AFB are being studied, mainly based on essential oils.

**Keywords:** *Paenibacillus larvae*, essential oils, quorum sensing, American foulbrood, *Apis mellifera*

### **1. Introduction**

Along with wild bees, honeybees are the most important crop pollinators [1, 2]. *Apis mellifera* pollinates 77% of the plants responsible for producing food resources which sustain the global human population [1]. Since 1998, individual beekeepers have reported the unusual weakening and mortality of colonies, particularly in France, Belgium, Switzerland, Germany, the United Kingdom, the Netherlands, Italy, Spain, and North America [3, 4]. Most scientists agree that there is no single explanation for the extensive colony losses, but that interactions between different stressors are likely involved [5].

American foulbrood (AFB) is the most severe bacterial disease that affects honey bees, having a nearly cosmopolitan distribution (**Figure 1**) [6]. AFB only kills

**Figure 1.** *Distribution of American foulbrood. (https://www.cabi.org/isc/datasheet/78183).*

infected honey bee larvae; however, it eventually leads to the collapse of the entire colony when left untreated. AFB is considered to be very contagious; therefore, it is a notifiable disease in most countries [7]. AFB's causative agent is *Paenibacillus larvae*, which is a flagellated gram-positive bacterium, whose main characteristic is the formation of highly resistant endospores. This pathogen affects the breeding during the larval or pupal stages [8]; its spores being the infectious form. Honey bee larvae are more susceptible to infection during the first 36 h after egg hatching [9], indeed only 10 spores are required to make a larva of less than 24 h old ill [10]. However, at later larval developmental stages, spore doses needed to successfully infect a larva are too high to occur under natural conditions [11]. The infection takes place by the ingestion of the spores with the food provided by adult worker bees (nurses) to the larvae [12]. The spores after germinating in the midgut of the larvae proliferate for several days. After this, *P. larvae* reaches the peritrophic matrix, penetrates the epidermal cells, produces septicemia causing death of the larva. Finally, dead larvae are digested by vegetative bacterial cells and converted to dry flakes containing millions of spores of *P. larvae* [12, 13]. The most evident symptoms of AFB are the irregular coating of the offspring, which show cells with cap and uncovered irregularly dispersed through the frames of the offspring; dark, sunken, and often perforated caps emitting a characteristic AFB odor; remnants of brown glue from the dead larvae forming a characteristic cord thread when removed with a wooden stick or an inlay; and a hard scale of larval residues at the bottom of the cell. The traditional diagnosis is made based on the observation of these clinical symptoms in the hive and in the microbial culture of material from infected colonies [14].

AFB was first described in South America (SA) in Argentina, in 1989, constituting the first sanitary challenge for beekeepers to overcome. It was hypothesized that the entrance of *P. larvae*, into the country was through bees imported from the USA [15]. AFB quickly spread to most important beekeeping centers of the country [16], with incidences as high as 30% in some geographic areas [17]. At least 30–45% of the colonies were lost due to AFB during those years (Eguaras, unpublished data). AFB was extended to Chile in 2002 and was controlled. New outbreaks were detected in 2005 in different regions [18] (**Table 1**).

In some countries the use of antibiotics, particularly tetracycline hydrochloride (OTC) [6, 12], is the most common method for prevention and treatment of

**27**

**1.1 Essential oils**

**Table 1.**

*Prevention and Control of American Foulbrood in South America with Essential Oils: Review*

Argentina Restricted distribution [15, 84] Bolivia No information available [84] Brazil Present [84] Chile Present [84] Colombia Disease never reported [84] Ecuador Disease never reported [84] French Guiana Disease never reported [84] Peru Disease not reported [84] Uruguay Present [84] Venezuela Disease never reported [84]

infected colonies. However, in most European countries the use of antibiotics is banned, since their use is known to generate several problems including the presence of chemical residues in the beehive products (honey, pollen and wax), which eventually may even affect consumer health. Moreover, antibiotic application can affect life of bees and can increase the risk of occurrence of resistant strains [19]. To date, the presence of OTC resistant strains has been reported in Argentina, United

Prevention and control measures of AFB in SA countries generally include vigilance for early diagnosis, isolation of apiaries with cases of AFB, and multiplication of healthy colonies with hygienic queens, among others [21]. Brazilian, Chilean, and Uruguayan authorities specifically recommend the burning of colonies containing clinical signs of the disease in order to control the outbreaks [21]. The use of antibiotics in SA is not allowed, except in Argentina [18]. The extensive use of OTC in this country has led to the development of resistant *P. larvae* isolates [16], which is a major concern for Argentine beekeepers. In contrast, in Uruguay and Chile, where their use is not authorized, no resistant strains have been detected [22]. The endospore resistance of *P. larvae* is an important problem in the control and prevention of AFB because these individuals can survive for more than 35 years in honey and/or beekeeping material and is resistant to high temperatures as well as to the most used disinfectants [10]. Most treatments are based on the use of broad spectrum antibiotics, which, in most cases, have been used continuously and excessively. In fact, different antibiotics, such as sulfathiazole and OTC, are able to inhibit the growth of *P. larvae*, but its use and abuse during the last years has led to the appearance of resistant strains and residues that contaminate the products of the hive. For these reasons, the use of antibiotics for the treatment and prevention of AFB is prohibited

in several countries, and the affected colonies must be destroyed [23].

essential oils [25–27], probiotics and propolis [28].

In this context, the development of alternative and effective methods for the control and prevention of AFB disease is crucial. These methods may consider the evidence of the bacteria-resistant phenomenon and meet the strict EU standards, as well as current trends in green consumption [24, 25]. Currently, alternative strategies for the prevention and treatment of AFB are being studied, mainly based on

In light of developments in the scientific field, the medicinal properties of plants

have received great interest due to their low toxicity, pharmacological activities

States, Italy, New Zealand and United Kingdom [16, 20].

*DOI: http://dx.doi.org/10.5772/intechopen.85776*

*Distribution of P. larvae in South America.*

*Prevention and Control of American Foulbrood in South America with Essential Oils: Review DOI: http://dx.doi.org/10.5772/intechopen.85776*


#### **Table 1.**

*Beekeeping - New Challenges*

**Figure 1.**

infected honey bee larvae; however, it eventually leads to the collapse of the entire colony when left untreated. AFB is considered to be very contagious; therefore, it is a notifiable disease in most countries [7]. AFB's causative agent is *Paenibacillus larvae*, which is a flagellated gram-positive bacterium, whose main characteristic is the formation of highly resistant endospores. This pathogen affects the breeding during the larval or pupal stages [8]; its spores being the infectious form. Honey bee larvae are more susceptible to infection during the first 36 h after egg hatching [9], indeed only 10 spores are required to make a larva of less than 24 h old ill [10]. However, at later larval developmental stages, spore doses needed to successfully infect a larva are too high to occur under natural conditions [11]. The infection takes place by the ingestion of the spores with the food provided by adult worker bees (nurses) to the larvae [12]. The spores after germinating in the midgut of the larvae proliferate for several days. After this, *P. larvae* reaches the peritrophic matrix, penetrates the epidermal cells, produces septicemia causing death of the larva. Finally, dead larvae are digested by vegetative bacterial cells and converted to dry flakes containing millions of spores of *P. larvae* [12, 13]. The most evident symptoms of AFB are the irregular coating of the offspring, which show cells with cap and uncovered irregularly dispersed through the frames of the offspring; dark, sunken, and often perforated caps emitting a characteristic AFB odor; remnants of brown glue from the dead larvae forming a characteristic cord thread when removed with a wooden stick or an inlay; and a hard scale of larval residues at the bottom of the cell. The traditional diagnosis is made based on the observation of these clinical symptoms in

*Distribution of American foulbrood. (https://www.cabi.org/isc/datasheet/78183).*

the hive and in the microbial culture of material from infected colonies [14].

detected in 2005 in different regions [18] (**Table 1**).

AFB was first described in South America (SA) in Argentina, in 1989, constituting the first sanitary challenge for beekeepers to overcome. It was hypothesized that the entrance of *P. larvae*, into the country was through bees imported from the USA [15]. AFB quickly spread to most important beekeeping centers of the country [16], with incidences as high as 30% in some geographic areas [17]. At least 30–45% of the colonies were lost due to AFB during those years (Eguaras, unpublished data). AFB was extended to Chile in 2002 and was controlled. New outbreaks were

In some countries the use of antibiotics, particularly tetracycline hydrochloride (OTC) [6, 12], is the most common method for prevention and treatment of

**26**

*Distribution of P. larvae in South America.*

infected colonies. However, in most European countries the use of antibiotics is banned, since their use is known to generate several problems including the presence of chemical residues in the beehive products (honey, pollen and wax), which eventually may even affect consumer health. Moreover, antibiotic application can affect life of bees and can increase the risk of occurrence of resistant strains [19]. To date, the presence of OTC resistant strains has been reported in Argentina, United States, Italy, New Zealand and United Kingdom [16, 20].

Prevention and control measures of AFB in SA countries generally include vigilance for early diagnosis, isolation of apiaries with cases of AFB, and multiplication of healthy colonies with hygienic queens, among others [21]. Brazilian, Chilean, and Uruguayan authorities specifically recommend the burning of colonies containing clinical signs of the disease in order to control the outbreaks [21]. The use of antibiotics in SA is not allowed, except in Argentina [18]. The extensive use of OTC in this country has led to the development of resistant *P. larvae* isolates [16], which is a major concern for Argentine beekeepers. In contrast, in Uruguay and Chile, where their use is not authorized, no resistant strains have been detected [22]. The endospore resistance of *P. larvae* is an important problem in the control and prevention of AFB because these individuals can survive for more than 35 years in honey and/or beekeeping material and is resistant to high temperatures as well as to the most used disinfectants [10]. Most treatments are based on the use of broad spectrum antibiotics, which, in most cases, have been used continuously and excessively. In fact, different antibiotics, such as sulfathiazole and OTC, are able to inhibit the growth of *P. larvae*, but its use and abuse during the last years has led to the appearance of resistant strains and residues that contaminate the products of the hive. For these reasons, the use of antibiotics for the treatment and prevention of AFB is prohibited in several countries, and the affected colonies must be destroyed [23].

In this context, the development of alternative and effective methods for the control and prevention of AFB disease is crucial. These methods may consider the evidence of the bacteria-resistant phenomenon and meet the strict EU standards, as well as current trends in green consumption [24, 25]. Currently, alternative strategies for the prevention and treatment of AFB are being studied, mainly based on essential oils [25–27], probiotics and propolis [28].

#### **1.1 Essential oils**

In light of developments in the scientific field, the medicinal properties of plants have received great interest due to their low toxicity, pharmacological activities

and economic viability [29]. These studies have focused on the benefits of phytochemicals extracted from plants and their effect on human health. The additives naturally obtained from plants can be individual compounds, groups of compounds or essential oils (EOs). In recent times, there has been an increase in the interest of the food industry in natural compounds, either by direct addition or by its use in synergy with other compounds. It has been reported that the direct addition of essential oils and extracts of aromatic plants to food products exerts its antioxidant or antimicrobial effect [30].

Plants and other natural sources can provide a wide variety of complex and structurally diverse compounds. Plant extracts and essential oils have antifungal, antibacterial and antiviral properties and have been evaluated worldwide as potential sources of new antimicrobial compounds, agents that promote food preservation and alternatives to treat infectious diseases [31, 32]. It has been reported that essential oils possess significant antiseptic, antibacterial, antiviral, antioxidant, antiparasitic, antifungal, and insecticidal activities [33, 34]. Therefore, essential oils can serve as powerful tools to reduce bacterial resistance [33]. Oily aromatic liquids called essential oils (also called volatile oils) are obtained from plant materials (leaves, buds, fruits, flowers, herbs, branches, bark, wood, roots and seeds).

Being natural mixtures of very complex nature, the essential oils can consist of approximately 20–60 components at quite different concentrations. Essential oils are characterized by two or three main components that are present in fairly high concentrations (20–70%) compared to other components that are present in trace amounts. The amount of different components of essential oils varies between different parts of plants and different plant species since they are derived chemically from terpenes and their oxygenated derivatives, i.e., terpenoids which are esters of aromatic and aliphatic acid, and phenolic compounds. An important characteristic of essential oils and their components is their hydrophobicity, which allows them to interact with the lipids present in the cell membrane of bacteria and mitochondria, making them more permeable by altering their cellular structures. This eventually results in the death of bacterial cells due to the leakage of critical molecules and ions from the bacterial cell. Some compounds modulate drug resistance by targeting efflux mechanisms in several species of gram-negative bacteria [35]. An important function of essential oils in nature is the protection of plants by acting as antifungal, antibacterial, antiviral and insecticidal agents and also protection against herbivores by reducing the appetite of herbivores for plants with such properties. Health Services and Human Services Public Health Services have recognized essential oils as safe substances, and some of them contain compounds that can be used as antibacterial additives [33]. The efficacy of EOs has been reported in several studies against pathogens and food contaminants [36], suggesting their applications in the food industry [34, 37]. Several EOs have been evaluated for the *in vitro* and *in vivo* control of *P. larvae* (**Table 2**), as well as their acute oral toxicity to *Apis mellifera* (**Table 3**).

### *1.1.1 In vitro assays to control P. larvae*

EOs from *Achyrocline satureioides*, *Carum carvi*, *Cinnamomum* spp., *Cinnamomum zeylanicum*, *Citrus paradise*, *Cuminum cyminum*, *Cymbopogon citratus*, *Eucalyptus cinerea*, *Melaleuca alternifolia*, *Mentha piperita*, *Minthostachys verticillata*, *Origanum majorana*, *Origanum vulgare*, *Polygonum bistorta*, *Salvia officinalis*, *Salvia sclarea*, *Syzygium aromaticum*, *Tagetes minuta*, *Thymus vulgaris*, *Verbena*, *Pimenta dioica* (L.) Merr., *Litsea cubeba* Pers., *Trachyspermum ammi* L., *Mentha arvensis* L., *Mentha spicata* L., *Illicium verum* Hook.f, *Myristica fragrans* Gronov., *Cinnamomum camphora* (L.) J. Presl., *Ocimum tenuiflorum* L., *Daucus carota* L., *Zingiber officinale* Rosc., and

**29**

**Essential oil** *Acantholippia seriphioides* A. Gray

*Achyrocline satureioides* Lam.

*Artemisia absinthium* L.

*Artemisia annua* L.

*Aloysia polystachia* Griseb.

*Carapa guianensis* Aubl.

*Carum carvi* L.

*Chamomilla recutita* L.

*Cinnamomum aromaticum* L.

*Cinnamomum camphora* (L) J. Presl.

*Cinnamomum glandulifera* Nees.

Broth macrodilution Broth macrodilution Agar diffusion Broth microdilution Broth microdilution Broth microdilution

Broth

Inhibitory

microdilution

Agar diffusion Agar diffusion Agar diffusion Agar diffusion Agar diffusion

Broth

Inhibitory

3200–0.78

286.2 ± 27.9 μg/ml

700 μg/ml

375.0 ± 34.8

[36]

[40]

microdilution

Agar dilution

Inhibitory

Inhibitory

10 μl

Inhibitory

0.015% (v/v) (strong

activity)

Inhibitory

10 μl

Noninhibitory

5, 10 μl

Inhibitory

5, 10 μl

Inhibitory

Inhibitory

Inhibitory Inhibitory

416 mg/L 402 mg/L 700–800 mg/L

25% (v/v)

900 mg/L

[42]

624 mg/L

[42]

647 mg/L

[42]

10 μl

Inhibitory

Inhibitory

**Technique**

**Activity**

**Amount tested**

**MICa** 236 mg/L 300 mg/L

**MBCb**

**References**

[26] [41]

*Prevention and Control of American Foulbrood in South America with Essential Oils: Review*

[27]

[35]

[35]

[34]

[34]

[36]

*DOI: http://dx.doi.org/10.5772/intechopen.85776*

[33]


*Prevention and Control of American Foulbrood in South America with Essential Oils: Review DOI: http://dx.doi.org/10.5772/intechopen.85776*

*Beekeeping - New Challenges*

or antimicrobial effect [30].

and economic viability [29]. These studies have focused on the benefits of phytochemicals extracted from plants and their effect on human health. The additives naturally obtained from plants can be individual compounds, groups of compounds or essential oils (EOs). In recent times, there has been an increase in the interest of the food industry in natural compounds, either by direct addition or by its use in synergy with other compounds. It has been reported that the direct addition of essential oils and extracts of aromatic plants to food products exerts its antioxidant

Plants and other natural sources can provide a wide variety of complex and structurally diverse compounds. Plant extracts and essential oils have antifungal, antibacterial and antiviral properties and have been evaluated worldwide as potential sources of new antimicrobial compounds, agents that promote food preservation and alternatives to treat infectious diseases [31, 32]. It has been reported that essential oils possess significant antiseptic, antibacterial, antiviral, antioxidant, antiparasitic, antifungal, and insecticidal activities [33, 34]. Therefore, essential oils can serve as powerful tools to reduce bacterial resistance [33]. Oily aromatic liquids called essential oils (also called volatile oils) are obtained from plant materials (leaves, buds, fruits, flowers, herbs, branches, bark, wood, roots and seeds).

Being natural mixtures of very complex nature, the essential oils can consist of approximately 20–60 components at quite different concentrations. Essential oils are characterized by two or three main components that are present in fairly high concentrations (20–70%) compared to other components that are present in trace amounts. The amount of different components of essential oils varies between different parts of plants and different plant species since they are derived chemically from terpenes and their oxygenated derivatives, i.e., terpenoids which are esters of aromatic and aliphatic acid, and phenolic compounds. An important characteristic of essential oils and their components is their hydrophobicity, which allows them to interact with the lipids present in the cell membrane of bacteria and mitochondria, making them more permeable by altering their cellular structures. This eventually results in the death of bacterial cells due to the leakage of critical molecules and ions from the bacterial cell. Some compounds modulate drug resistance by targeting efflux mechanisms in several species of gram-negative bacteria [35]. An important function of essential oils in nature is the protection of plants by acting as antifungal, antibacterial, antiviral and insecticidal agents and also protection against herbivores by reducing the appetite of herbivores for plants with such properties. Health Services and Human Services Public Health Services have recognized essential oils as safe substances, and some of them contain

compounds that can be used as antibacterial additives [33]. The efficacy of EOs has been reported in several studies against pathogens and food contaminants [36], suggesting their applications in the food industry [34, 37]. Several EOs have been evaluated for the *in vitro* and *in vivo* control of *P. larvae* (**Table 2**), as well as their

EOs from *Achyrocline satureioides*, *Carum carvi*, *Cinnamomum* spp., *Cinnamomum* 

*zeylanicum*, *Citrus paradise*, *Cuminum cyminum*, *Cymbopogon citratus*, *Eucalyptus cinerea*, *Melaleuca alternifolia*, *Mentha piperita*, *Minthostachys verticillata*, *Origanum majorana*, *Origanum vulgare*, *Polygonum bistorta*, *Salvia officinalis*, *Salvia sclarea*, *Syzygium aromaticum*, *Tagetes minuta*, *Thymus vulgaris*, *Verbena*, *Pimenta dioica* (L.) Merr., *Litsea cubeba* Pers., *Trachyspermum ammi* L., *Mentha arvensis* L., *Mentha spicata* L., *Illicium verum* Hook.f, *Myristica fragrans* Gronov., *Cinnamomum camphora* (L.) J. Presl., *Ocimum tenuiflorum* L., *Daucus carota* L., *Zingiber officinale* Rosc., and

acute oral toxicity to *Apis mellifera* (**Table 3**).

*1.1.1 In vitro assays to control P. larvae*

**28**


**31**

**Essential oil** *Cymbopogon martini* Stapf.

*Cymbopogon nardus* L.

*Daucus carota* L.

*Eucalyptus cinerea* F. Muell

*Eugenia* spp. *Illicium verum* Hook.f.

*Illicium verum* Hook.f.

*Lavandula officinalis* L.

*Laurus nobilis* L.

*Lepechinia floribunda* Benth.

*Lippia turbinata* Griseb

*Litsea cubeba* Pers.

*Litsea cubeba* Pers.

**Technique** Broth microdilution Broth microdilution Agar diffusion Broth microdilution Agar diffusion Agar diffusion Agar diffusion Broth microdilution

Broth

Inhibitory

macrodilution

Broth

Inhibitory

microdilution

Broth

Inhibitory

microdilution

Broth

Inhibitory

microdilution

Broth

Inhibitory

macrodilution

Agar diffusion

Broth

Inhibitory

3200–0.78 μg/

85.0 ± 7.9 μg/ml

186.0 ± 21.2 μg/

[36]

ml

ml

microdilution

Inhibitory

10 μl

Inhibitory Inhibitory

3200–0.78

278.6 ± 21.2 μg/ml

350–400 μg/ml

1000 μg/ml 12,879 μg/ml

394 mg/L 866 mg/L

518 mg/L

[26]

[26]

[36]

365.0 ± 32.1

10 μl

Inhibitory

Inhibitory

10 μl

Inhibitory Inhibitory

3200–0.78 μg/

412.8 ± 26.0 μg/ml

589.6 ± 48.2 μg/ml

[36] [33] [32] [36]

ml

10 μl

Inhibitory

Inhibitory

**Activity**

**Amount tested**

**MICa** 1195 mg/L 319 mg/L

595 mg/L

[42]

1208 mg/L

[42]

**MBCb**

**References**

*Prevention and Control of American Foulbrood in South America with Essential Oils: Review*

[36]

[45]

[39]

[36]

*DOI: http://dx.doi.org/10.5772/intechopen.85776*

[36]

*Beekeeping - New Challenges*


*Prevention and Control of American Foulbrood in South America with Essential Oils: Review DOI: http://dx.doi.org/10.5772/intechopen.85776*

*Beekeeping - New Challenges*

**30**

**Essential oil** *Cinnamomum zeylanicum* L.

**Technique** Agar diffusion

Broth

Inhibitory Inhibitory Inhibitory Inhibitory

*Cinnamomum zeylanicum* + *Thymus vulgaris* L.

*Citrus limon* L.

*Cinnamomum zeylanicum* + *Thymus vulgaris* L.

*Citrus limon* L.

*Citrus nobilis* Lour

*Citrus reticulata* var. *madurensis* Blanco

*Copaifera officinalis* L.

*Copaifera officinalis* L. nanoemulsion

*Cymbopogon citratus* + *Thymus vulgaris* L.

*C. citratus* + *T. vulgaris* + *Satureja hortensis* L. + *Origanum* 

*vulgare* L. + *Ocimum basilicum* L.

*C. citratus* + *T. vulgaris* + *O. basilicum*

Agar dilution Agar dilution Agar dilution

Inhibitory

Inhibitory

Inhibitory

Broth

Inhibitory

macrodilution

Broth

Inhibitory

microdilution

Broth

Inhibitory

macrodilution

Broth

Inhibitory

microdilution

Broth

Inhibitory

microdilution

Agar diffusion

Agar dilution

Broth

Inhibitory Inhibitory

microdilution

Inhibitory

0.12–1.0% (v/v)

1.56% (v/v) 0.39% (v/v) 25–100 μg/ml 25–175 μg/ml 50–350 μg/ml

Inhibitory

10 μl

58–83 μg/ml 25–100 mg/L 38–50 μg/ml 25–67 μg/ml

66.6 μg/ml

764 mg/L 66.6 μg/ml

764 mg/L 815 mg/L

2447 mg/L

[42]

[34]

[34]

[27]

[50]

[40]

[40]

[40]

2293 mg/L

[42]

95.83 μg/ml

[42]

2293 mg/L

[26]

95.83 μg/ml

[42]

108–112 μg/ml

25–100 mg/L

[45]

[46]

[6]

[42]

macrodilution

Inhibitory

2 mg/ml

**Activity**

**Amount** 

**MICa**

**MBCb**

**References**

[36, 59]

**tested**


**33**

**Essential oil** *Ocimum tenuiflorum* L.

*Pimenta dioica* (L.) Merr.

*Pimpinella anisum* L.

*Salvia officinalis* L.

*Salvia sclarea* L.

*Satureja odora* Griseb.

*Schinus molle* L.

*Syzygium aromaticum* L.

*Syzygium aromaticum* L.

*Syzygium aromaticum* L.

**Technique** Agar diffusion Broth microdilution Agar diffusion Broth microdilution Agar diffusion

Inhibitory Inhibitory Inhibitory

300 μg/ml

Broth macrodilution Agar diffusion

Inhibitory Inhibitory

> Agar diffusion

Agar dilution

Broth

Inhibitory

microdilution

Broth

Inhibitory

macrodilution

Agar diffusion Agar diffusion Agar diffusion

Agar dilution

Inhibitory

0.015% (v/v) (strong

activity)

Inhibitory

10 μl

Inhibitory

5 μl

Inhibitory

10 μl

Inhibitory

0.06% (v/v) (strong

activity)

700–800 mg/L

666 mg/L

900 mg/L

[42]

[42]

[34]

[35]

[35]

[34]

Inhibitory

10 μl

10 μl

5 μl

10 μl

5 μl

Inhibitory Inhibitory

3200–0.78 μg/

78.0 ± 8.2 μg/ml

162.0 ± 18.2 μg/

ml

ml

10 μl

Inhibitory Inhibitory

3200–0.78

412.8 ± 26.0 μg/ml

589.6 ± 48.2

[36]

10 μl

**Activity**

**Amount tested**

**MICa**

**MBCb**

**References**

[36]

*Prevention and Control of American Foulbrood in South America with Essential Oils: Review*

[35]

[35]

[34]

[34]

*DOI: http://dx.doi.org/10.5772/intechopen.85776*

[36] [35] [35] [46]

[36]

#### *Beekeeping - New Challenges*


#### *Prevention and Control of American Foulbrood in South America with Essential Oils: Review DOI: http://dx.doi.org/10.5772/intechopen.85776*

*Beekeeping - New Challenges*

[34]

**32**

**Essential oil** *Melaleuca alternifolia* Maiden & Betche

**Technique** Agar diffusion

Agar dilution

Broth

Inhibitory Inhibitory Inhibitory

Broth

Inhibitory

microdilution

Broth

Inhibitory

microdilution

Broth

Inhibitory

microdilution

Broth

Inhibitory

microdilution

Agar diffusion

Broth

Inhibitory

3200 to

145.6 ± 15.4 μg/ml

775 mg/L

256.0 ± 26.5 μg/

[36]

[42]

[33]

[36]

ml

0.78 μg/ml

microdilution

Broth

Inhibitory

macrodilution

Agar diffusion Agar diffusion

Broth

Inhibitory

3200–0.78

285.8 ± 29.2 μg/ml

350–450 μg/ml

0.06–0.12% (v/v)

371.3 ± 29.0

[36]

[40]

[34]

microdilution

Agar dilution

Inhibitory Inhibitory

Inhibitory

10 μl

Inhibitory

10 μl

Inhibitory

10 μl

*Mentha arvensis* L.

*Mentha* (hybrid)

*Mentha rotundifolia* L.

*Mentha spicata* L.

*Minthostachys mollis* Kunth.

*Minthostachys verticillata* Griseb

*Myristica fragrans* Gronov.

*Myristica fragrans* Gronov.

*Ocimum basilicum* L.

microdilution

Inhibitory

0.015–0.12% (v/v) (strong

activity)

1095 mg/L 0.18–1.5% (v/v)

331 mg/L 1000–1800 μg/ml

144.7 ± 17.2 μg/ml

600–700 μg/ml

600–1000 μg/ml

1600 ≥ 2000 μg/

[81]

[36]

ml

1000–1200 μg/ml

[81]

248.0 ± 23.4 μg/

[36]

ml

585 mg/L 1600–2000

[81]

[42]

1187 mg/L

[42]

[27]

Inhibitory

10 μl

**Activity**

**Amount** 

**MICa**

**MBCb**

**References**

[34]

**tested**

