Blackleg: A deadly disease with a hidden cause

Abstract

Blackleg is an infectious disease that mainly affects cattle and rarely affects other ruminants. It is characterized by hemorrhagic blackleg myositis. Clostridium chauvoei is a highly pathogenic anaerobic, endospore-forming Gram- positive bacteria that causes blackleg disease. Blackleg disease was first reported in 1870, but the causative bacterium C. chauvoei was not described until 1887. Clostridium chauvoei is found in grassland, fresh water, silage, soil, and the excrement of healthy animals. Cattle intestines and pasture soil have both been found to contain C. chauvoei spores, suggesting that the infection is contracted by ingesting the spores. Significant lesions are less common in the heart and more common in the skeletal muscle. Anaerobic culture, polymerase chain reaction, immunodetection using fluorescent antibody tests, and immunohistochemistry are methods for identifying bacteria. The production of gas in animal tissues, known as emphysematous swelling, is a hallmark of blackleg illness. Swelling usually affects big muscles including the neck, shoulders, thighs, and hips. Most incidences of blackleg illness in cattle occur during the warmer months of the year, and the disease typically manifests seasonally. Penicillin treatment and surgical removal of the lesions, including fasciotomy, are recommended for infected animals that are not in danger of death. Vaccination is a crucial part of the health management of many livestock production enterprises and can prevent blackleg illness. Blackleg disease can be controlled by destroying the carcass and burning the top layer of soil to eradicate any remaining spores.

Introduction

Blackleg is an infectious disease that mainly affects cattle and rarely affects other ruminants. The condition is characterized by hemorrhagic blackleg myositis (Abreu et al., 2018). This illness is caused by the anaerobic Gram-positive bacillus Clostridium chauvoei, which thrives in the soil as hardy spores (Morrell et al., 2022). Blackleg disease was first reported in 1870, but the causative bacterium C. chauvoei was not described until 1887 (Sousa et al., 2024). The spores of C. chauvoei can endure for years in the soil. Furthermore, they are impervious to chemical disinfectants, heat, cold, drought, and UV light (Rychener et al., 2017). Animals consume spores through their diet, which are believed to travel through the digestive system, enter the bloodstream, and then go to different muscles and organs where they stay until they are aroused to cause illness (Bagge et al., 2009).

Spores of C. chauvoei can develop into active bacteria in an animal host, producing a strong exotoxin that damages local tissue and frequently results in toxemia (Salvarani and Vieira, 2024). The production of gas in animal tissues, known as emphysematous swelling, is a hallmark of blackleg illness (Santos et al., 2019). Swelling usually affects big muscles including the neck, shoulders, thighs, and hips (Tagesu et al., 2019). The swelling feels painful and hot. Furthermore, infections that frequently affect ruminants are typified by necrosis of the heart and striated muscle, which frequently results in unexpected death (Sousa et al., 2024). Gas can be felt beneath the skin if the bulge is compressed. Animals usually develop symptoms within 12 to 48 hours. Animals are typically found dead without previously showing signs of illness (Hansford, 2020).

The prevalence of blackleg disease in ruminants has decreased dramatically due to the use of available and efficient vaccinations and antibiotics (Guizelini et al., 2020). Blacklegs are most common in young cattle between the ages of 6 months and 2 years. This is because colostrum does not provide young cows with enough passive immunity, especially calves from heifers (Ayele et al., 2016). Furthermore, cattle older than 2 years are rarely affected by the disease, most likely as a result of protection brought up by vaccination or natural exposure (Heckler et al., 2018). Nonetheless, some occurrences do happen in calves older than 2 years, and they are frequently linked to frequent syringe usage (Ayele et al., 2016). There is no direct contact between the sick and healthy animals that spread this disease.

This severe infectious disease affects ruminants worldwide and results in large financial losses for the cattle industry (Nampanya et al., 2019). Blackleg is a persistent risk to livestock that have not been vaccinated or who have received subpar vaccination due to the prevalence of C. chauvoei spores in the soil (Morrell et al., 2022). As a result, outbreaks of this disease are more common in livestock whose immunization has been ignored, but they nonetheless occasionally occur in vaccinated livestock. This review article aims to provide an overview of current knowledge regarding blackleg disease, including its causes, symptoms, diagnosis, prevention, and treatment.

Etiology

Clostridium chauvoei is a highly pathogenic, anaerobic, endospore-forming Gram-positive bacteria that causes blackleg disease. It requires an enriched medium to develop and generate endospores that resemble lemons (Frey and Falquet, 2015). In 2013, the first draft genome sequence of a virulent strain of C. chauvoei was made public; it had 2.8 million base pairs (bp) (Falquet et al., 2013). Furthermore, the bacteria contained cryptic plasmids that were approximately 5.5 kbps in size. (Frey et al., 2012). The whole genome sequences of 20C. chauvoei strains that were isolated over a 64-year span from four different continents have been identified and examined. The strains examined in this study are highly conserved, according to the study’s findings, which further implies that C. chauvoei evolution has come to a standstill (Rychener et al., 2017).

The very tiny genome of C. chauvoei (4.2 million bp) compared with other Clostridium species, including Clostridioides difficile, indicates that it has adapted to a restricted host range (sheep, goats, and cattle), where it can multiply and spread illness (Schüler et al., 2024). Nevertheless, a comparison of the circular genome sequences of the field strain 12S0467 isolated in Germany and the type strain of C. chauvoei American Type Culture Collection (ATCC) 10092 showed new differences in regulatory genes, suggesting that C. chauvoei has unique control over regulatory events in contrast to other Clostridium species (Thomas et al., 2017).

The 69 genes in C. chauvoei encode sporulation and dormancy mechanisms, which are virulence factors that enable the pathogen to endure harsh environmental conditions and maintain its potential for infection for years (Frey and Falquet, 2015). Genes in the Clostridia cluster I group, which include Clostridium novyi, C. botulinum, C. septicum, C. perfringens, C. tetani, C. haemolyticum, and have been found to be identical to those in C. chauvoei that are involved in sporulation and germination (Thomas et al., 2017). Additionally, C. chauvoei produces a number of soluble antigens linked to pathogenicity as well as cellular antigens (such as flagella and somatic) (Sousa et al., 2024).

Cellular antigens

Somatic antigens are found in C. chauvoei bacterial cells. Since the antigen is thought to be a crucial immunogenic element that protects against C. chauvoei infection, both the past and present vaccines either contain bacterin (Hamzavipour et al., 2024). Flagellin, which is encoded by the fliC sequence, is the most well-studied of the flagellar antigens (Razim et al., 2021). The pathogen-associated molecular patterns observed in flagellin are identified by monocyte and fibroblast-expressed toll-like receptor 5 (Jang et al., 2015). Protein secretion is activated when conserved portions of flagellin (the N and C termini) are bound by receptors on the surface of viscoelastic animal tissue cells (Willing et al., 2015).

Studies have been conducted to investigate the use of recombinant flagellin supermolecules to describe and assess its protective action. According to research, mice that receive recombinant flagellin have inadequate protective immunity, suggesting that conformation- dependent epitopes are essential for the development of immunity against blackleg illness (Zou et al., 2021). It has frequently been suggested that the low protective action of previously identified recombinant flagellin supermolecules was caused by the researchers’ failure to take into account the presence of at least two copies of the fliC sequence in the body of C. chauvoei (Jabbari et al., 2015). Furthermore, the majority of the strains under investigation had three copies of the flagellin cistron variants fliC1, fliC2, and fliC3, showing 91.8% organic compound identity within a single strain and 82%–96% identity between paralogs of different strains (Kojima et al., 2000). In a different investigation, three fliC genes were also identified (Sasaki et al., 2002). There has been no investigation of C. chauvoei cell surface-associated antigens other than flagellin. There are a number of significant cell surface proteins linked to C. chauvoei that exhibit protective antigenicity against other bacteria, including ribosomal supermolecule L10, enolase, chaperonin, flavoproteins, and glycosyl hydrolases (Frey et al., 2012). To determine how these surface-associated proteins contribute to the protection against blackleg disease, more investigation is necessary.

Soluble antigens and toxins

The pathophysiology of blackleg illness is significantly influenced by toxins, which are soluble antigens. The current list of five recognized C. chauvoei toxins includes the hemolytic leukocidin CctA, oxygen-insensitive hemolysin D (or hemolysin III), DNase (ß-toxin), hyaluronidase Nag (formerly known as γ-toxin), and neuraminidase/sialidase NanA (Dangi et al., 2017). The enzyme DNase (β-toxin), a deoxyribonuclease- type protein that is thermostatically responsible for the breakdown of muscle cell nuclei, contributes to clostridial myonecrosis (Los et al., 2000).

It has been identified in >80% of C. chauvoei strains recovered from cattle; however, these strains exhibit variable toxin-producing capacity (Ziech et al., 2018). Two genes encoding the large and small subunits of exo-deoxyribonuclease VII were identified in whole genome investigations of C. chauvoei. These genes most likely reflect the DNase activity of C. chauvoei (Thomas et al., 2017). The exo-deoxyribonuclease VII gene is present in all 20 C. chauvoei strains (Daly et al., 2009). The heat-inactivated enzyme hyaluronidase (γ-toxin) breaks down hyaluronic acid, which is believed to be the cause of the breakdown of loose connective tissue that envelops muscle (Hynes and Walton, 2000). This mechanism allows C. chauvoei to proliferate throughout infected host tissue.

Strong hemolytic activity is exerted by the spore- forming, oxygen-stable hemolysin leukocidin C. chauvoei cytotoxin A (CctA), which is seen as a halo surrounding the colonies on blood agar growth medium (Ziech et al., 2018). The molecular mass of mature CctA is 32.2 kDa. The primary hemolysin and toxin that C. chauvoei produces, CctA, has been demonstrated to be extremely cytotoxic to the bovine epithelial cell line ECaNEp (Frey et al., 2012). Additionally, the effectiveness of a blackleg vaccine that contained purified recombinant CctA as the only antigen was tested using a traditional assay, and it shielded 80% of guinea pigs from virulent C. chauvoei exposure (Frey et al., 2012). CctA is a good candidate for a blackleg vaccine and for evaluating the efficacy of existing vaccinations since antibodies against it contribute significantly to the protective immunity provided against blackleg (Khiav and Zahmatkesh, 2021).

The previously identified oxygen-stable necrotic hemolysin (toxin-a) is most likely CctA, despite the fact that hemolysin toxin-a’s stated molecular mass is 25 kDa, which is much smaller (Hamzavipour et al., 2024). This toxin-a might be hemolysin III, also known as hemolysin D or toxin-d (protein #276), which is present in the genome of C. chauvoei and has a molecular mass of about 25 kDa (Nicholson et al., 2019). Because hemolysin III is expressed or carried by a wide range of pathogenic, commensal, and environmental gram- positive bacteria, it should be noted that it is not exclusive to C. chauvoei (Prajapati et al., 2024). The nature of hemolysin III in C. chauvoei is unclear, but it has been reported to resemble tetanolysin from C. tetani and the θ toxin from C. perfringens (Baldassi, 2005). The pathogen does not create any other entities with detectable hemolytic activity than CctA, as evidenced by the fact that the monospecific antibody utilized was directed against CctA and totally neutralized all hemolytic activity expressed by C. chauvoei (Frey et al., 2012).

The 81-kDa protein known as neruriminidase/sialidase (NanA) is secreted as a dimer and has been thoroughly described (Uchiyama et al., 2009). The nanA gene, which has been fully conserved in C. chauvoei strains identified over the course of 60 years from various geographic regions across four continents, encodes (Vilei et al., 2011). The sialidase activity of C. chauvoei was completely neutralized by a recombinant molecule of nanA with a sialic acid binding domain (CBM40) (Falquet et al., 2013). As a result, NanA can also serve as an antigen to support protective immunity.

History

Although blackleg disease, which affects cattle, sheep, and other ruminants, was initially identified in 1870, the bacteria that causes it, C. chauvoei, was not identified until 1887 (Sousa et al., 2024). There are 23 strains of the bacteria known as C. chauvoei, which were named in honor of French bacteriologist Professor J.A.B. Chauveau (Rychener et al., 2017). Vaccination is the most effective disease prevention strategy worldwide. The illness was initially discovered in Nigeria in 1929 by Anon, and it continues to be a significant cattle issue there (Useh et al., 2003). Although vaccination against this disease has been in place since 1930, outbreaks are nevertheless occasionally reported each year (Ziech et al., 2018). In Nigeria, this disease is categorized as a “List A disease” since it is linked to a high yearly livestock fatality rate in the nation (Useh et al., 2006). Given the high yearly mortality rate of ruminants from blackleg illness, the economic losses must be enormous, yet they have not been measured.

Epidemiology

Blackleg typically targets unvaccinated cattle between the ages of 6 months and 2 years who are in good nutritional condition (Ayele et al., 2016). However, isolated instances have been documented in fetuses, animals older than 2 years, and younger calves (Abreu et al., 2017; Heckler et al., 2018). In rare cases, blacklegs in vaccinated cattle have been documented (Abreu et al., 2018). Sheep rarely have blacklegs. In sheep, it is critical to differentiate between gas gangrene linked to external pathogenesis caused by C. chauvoei and endogenous pathogenesis caused by blackleg, particularly in cases where the infection portal of entry is unknown (Beigh et al., 2017). Other animal species, such as horses (Uzal et al., 2022), elephants (Miyashiro et al., 2007), chickens (Prukner-Radovcic et al., 1995), deer (Echenique et al., 2018), minks (Langford, 1970), and ostriches (Lublin et al., 1993), have also been reported to be infected by C. chauvoei. It remains mostly unclear what causes this infection in animals other than sheep and cattle. Humans, dogs, cats, and rabbits are believed to be immune to C. chauvoei infection (Sousa et al., 2024). Numerous nations, including Kazakhstan (Abutalip et al., 2023), the United States (Okafor et al., 2023), Iran (Espíndola et al., 2021), Austria (Wolf et al., 2017), Algeria (Gacem et al., 2015), Pakistan (Hussain et al., 2019), Ethiopia (Gedefa, 2021), Taiwan (Huang et al., 2013), Brazil (Heckler et al., 2018), Zambia (Hang’ombe et al., 2000), and Belarus (Blokhin et al., 2022), have reported cases involving animals. However, there are many more countries that do not register the disease. Clostridium chauvoei is found in grassland, fresh water, silage, soil, and the excrement of healthy animals (Bagge et al., 2009). It seems that the main way that animals become sick is through soil polluted with spores from the carcasses and excrement of diseased animals (Morrell et al., 2022). Although the ability of C. chauvoei to reproduce in soil has not been established, its spores can endure for extended periods of time there, and specific environmental factors, such as high temperatures and humidity, can help them survive (Bilska et al., 2024). The prevalence of blackleg disease is associated with the type of soil, amount of organic matter present, soil excavation, flooding, and yearly rainfall. This highlights the significance of these predisposing conditions in the transmission of C. chauvoei spores (Ziech et al., 2018). The majority of blackleg cases occur when animals grazing on contaminated pastures are relocated to new pastures following a period of intense rainfall. Less frequently, penned animals get blacklegs, most likely as a result of eating spore-tainted feed (Bagge et al., 2009). Rainfall and the incidence of blackleg disease in grazing animals are related because heavy rainfall induces anaerobic soil conditions that encourage bacterial growth and extensive spore spreading (Groseth et al., 2011).

Pathogenesis

There is considerable regarding agreement over the processes underlying the pathogenesis of C. chauvoei, although blackleg disease is a clinically recognized illness. Cattle intestines and pasture soil have both been found to contain C. chauvoei spores, suggesting that the infection is contracted by ingesting the spores (Ziech et al., 2018). Macrophages use Peyer’s patches to carry spores that are eaten or generated following a germination cycle in the bowel from intestinal or oral cavity lesions to muscles and tissues (Pires et al., 2012). Spores remain latent after reaching the tissue until specific circumstances, including anaerobiosis, occur, causing germination, multiplication, and subsequent generation of exotoxins (Abreu et al., 2018).

After traumatic damage, lactic acid concentrations rise anaerobically during glycolysis (the conversion of pyruvate to lactate) and muscle tissue oxygen levels fall, which promotes spore germination, bacterial growth, and the eventual synthesis of toxins (Pires et al., 2017). Nevertheless, this theory is insufficient to explain why the diaphragm or heart are the sole organs impacted at particular periods, or why only young animals are impacted. Furthermore, it’s unclear if circumstances let latent spores germinate in the absence of muscle damage. This could be because greater rates of muscle synthesis result in higher muscle glycogen concentrations, which could act as a substrate for C. chauvoei (Ziech et al., 2018). Healthy cattle corpses contain latent C. chauvoei spores in organs including the liver and spleen (Morrell et al., 2022). Clostridium chauvoei was detected by microbiological culture in 1.7% of liver samples and 7.5% of muscle samples from two slaughterhouses in Sao Paulo, Brazil, as part of a surveillance investigation (Kuhnert et al., 1997).

The significance of macrophages in the early pathophysiology of blackleg illness was supported by a recent study showing that both vegetative and sporulating forms of C. chauvoei could withstand the microbicidal effects of macrophages in mouse and bovine monocyte-derived macrophages (Pires et al., 2017). Following infection with vegetative C. chauvoei, the researchers observed proinflammatory cytokine patterns in bovine macrophages, including IL-12 and IL-23 transcription. On the other hand, an anti-inflammatory cytokine profile, including the induction of IL-10 and TGF-beta transcription, was noted in bovine macrophages infected with C. chauvoei spores (Pires et al., 2017). It is possible that the spores’ delay following macrophage internalization is due to the anti-inflammatory characteristic they produce. Future research should therefore examine the potential contribution of genetic predisposition to the onset of blackleg illness. The ability of phagocytic cells, particularly macrophages, to eliminate C. chauvoei following internalization can be genetically characterized as a starting point (Vilei et al., 2011). Figure 1 illustrates the interaction and processes involved in the pathogenesis of blackleg disease.

Fig. 1. Fig. 1. Pathogenic process of blacklegs in cattle.

Pathology

Typically, animals that pass away from blackleg illness are in very good nutritional condition (Abreu et al., 2018). Significant lesions are less common in the heart and more common in the skeletal muscle (Okafor et al., 2023). Other tissues, such as the thymus, intestine, meninges, liver, and brain are rarely affected. Only 3% of instances of this disease are thought to have heart abnormalities that are free of skeletal muscle injury (Heckler et al., 2018). Clostridium chauvoei lesions often target many pelvic and pectoral girdle muscle groups, but they can also affect any striated muscle, including the heart (Abreu et al., 2018). According to a retrospective analysis, the majority of the cases examined had heart and skeletal muscle lesions, despite other sources claiming that cardiac lesions linked to black leg illness are uncommon (Uzal et al., 2003). It is probable that, in some instances, at least, potential lesions in the heart and other skeletal muscles, such as the diaphragm, were missed because the carcasses were not examined further once lesions were discovered in the big skeletal muscles. This is a classical detection bias.

External examination may reveal an area of poorly defined swelling over the affected muscle area, where the skin is stretched and typically dark in color. Palpation may reveal crepitation caused by the buildup of subcutaneous serosanguineous fluid and emphysema (Abreu et al., 2018). In the center of the lesion, the affected muscle is dry and brittle, dark red to black, and edematous. It also has numerous tiny cavities formed by gas bubbles near the lesion’s periphery (Abreu et al., 2017). However, the bacteria’s synthesis of butyric acid is linked to the sweet, rancid butter-like smell emanating from damaged tissue (Sousa et al., 2024). The main lesions in heart-related instances are focally large black regions of bleeding, necrosis, and sporadic emphysema, which can occur anywhere in the myocardium (Okafor et al., 2023). Pericarditis is frequent, and different levels of fibrin frequently cling to the lining of the pericardial sac (Ziech et al., 2018). Furthermore, the pericardial sac frequently contains an excess of serosanguineous fluid (Morrell et al., 2022). Other sporadic lesions include pulmonary congestion, pulmonary edema, and fibrinohemorrhagic pleurisy, which mostly affect the pleura next to the heart (Hussain et al., 2019). Glossitis and necrotizing enteritis are uncommon.

Blackleg disease lesions resemble those in the skeletal and heart muscle under a microscope. Both tissues’ affected muscle fibers exhibit degenerative or necrotic alterations that include hypereosinophilia, vacuolization, swelling, fragmentation with loss of transverse stripes, and contraction bands (Abreu et al., 2018). Initially, neutrophils may enter the interstitium and broken fibers; later on in the infection process, macrophages, plasma cells, and lymphocytes may also infiltrate (Morrell et al., 2022). However, in the majority of blackleg disease cases, leukocyte infiltration is not a noticeable symptom (Blokhin et al., 2022). Hemorrhage and proteinaceous edema fluid along with many Gram-positive rods, some of which include subterminal spores, either alone or in clusters, increase the interstitium (Salvarani and Vieira, 2024). The interstitium of the skeletal muscle typically contains large, empty vacuoles made of gas bubbles, but the myocardium hardly ever does (Abreu et al., 2017). Fibrin thrombi are frequently found in vessels, and the arteries and arterioles may have fibrinoid necrosis accompanied by intramural neutrophil infiltration (Morrell et al., 2022). Vascular alterations in fibrinosuppurative pericarditis and pleuritis may resemble those reported in muscle (Abreu et al., 2018).

Diagnosis

The identification of C. chauvoei in afflicted tissues, clinical symptoms, and suitable gross and microscopic results are the foundation for a conclusive diagnosis of blackleg disease (Bagge et al., 2009). Anaerobic culture (Adib et al., 2023), polymerase chain reaction (PCR) (Sasaki et al., 2000), immunodetection with fluorescent antibody test (FAT) (Assis et al., 2007), and immunohistochemistry (IHC) (Casagrande et al., 2015) are methods for identifying bacteria. It is not always possible to isolate bacteria because of challenges in transporting samples to the lab and because not all diagnostic labs around the world have anaerobic culture facilities. The isolation of C. chauvoei may also be hampered by postmortem pollutants, such as other quickly developing anaerobes such as C. septicum (Halm et al., 2010).

When fresh tissue for microbiological culture is unavailable, FAT and IHC are particularly helpful methods. Both approaches have comparable sensitivity and specificity and are quick and easy to use. FAT can be used on smears or tissue that has been temporarily fixed in formalin, and it is quicker and less expensive than IHC (Assis et al., 2007). IHC is particularly beneficial for retrospective research by detecting organisms in lesions (Casagrande et al., 2015).

PCR is a very helpful technique for detecting C. chauvoei in pure cultures, clinical samples, or tissues that have been formalin-fixed and paraffin-embedded (FFPE) (Bagge et al., 2009). In an investigation, C. chauvoei was found in filter paper soaked with tissue samples collected from suspected blackleg cases using a PCR assay. This demonstrated that the proposed method is dependable, feasible for field settings, and economical (Espíndola et al., 2021). Multiplex real-time PCR has made it possible to identify C. chauvoei and C. septicum simultaneously, in addition to conventional PCR (Lange et al., 2010). To identify pathogens with high specificity, a recombinant C. chauvoei flagellin ELISA was created; however, this assay has not been utilized for routine testing, and more research is required to assess its diagnostic value (Usharani et al., 2015).

Differential diagnosis

Anthrax

The clinical symptom of an acute hemorrhagic disease like anthrax is abrupt mortality. Similar to blackleg disease, dark-colored fluid spilling from bodily orifices, enlarged spleen, lack of rigor mortis, and liver and kidney degeneration (Khairullah et al., 2024). Indeed, the resemblances are so great that diagnosis is frequently only possible upon laboratory identification of the particular bacterium.

Malignant edema

There are a few ways in which malignant edema is not the same as blackleg illness. The bacterium Clostridium septicum causes this illness (Nanjappa et al., 2015). Compared with blackleg sickness, this illness is more prevalent in older animals and is more likely to strike in winter. Amblyomma mites may be present in this instance of the condition, though, as evidenced by the presence of ehrlichia bodies in the endothelial cells of the blood capillaries in the brain and jugular veins (Drevets et al., 2004).

Snakebite

Snakebite is also one of the diseases that must be recognized from blackleg disease because this disease causes rapid death without clinical indications and is occasionally accompanied by edema in the pharynx and chest. It can be confused with sub-acute sickness (Lai et al., 2024). Bites to the head, neck, and snout often indicate acute systemic illness and death. Bite marks and typically local tissue swelling, redness, and bleeding may be seen upon viewing or biting, indicating snakebite envenomation (Mehta and Sashindran, 2002).

Clinical symptoms

Cattle

There is considerable lameness if the animal is seen before it dies, frequently accompanied by noticeable swelling in the affected leg’s upper portion (Tagesu et al., 2019). A deeper look reveals that the animal is extremely depressed, has rumen stasis, full anorexia, a high temperature of 41°C, and a heart rate of 100 to 120 beats per minute (Salvarani and Vieira, 2024). Pyrexia is not always present. Early on, the swelling is hot and unpleasant to the touch, but it quickly cools down and stops the pain. It is edematous and emphysematic, and when you press on it, it crackles like twisted tissue paper (Ziech et al., 2018).

Grunts were heard along with a gulp of breath. As the illness worsens, breathing becomes harder, the patient collapses, the temperature drops to normal or slightly below normal, and death occurs soon after. In rare, nonfatal situations, cattle develop severe abscesses that heal slowly, leaving them in a very weak, emaciated state (Ziech et al., 2018). Animals that are impacted in this way should be put down because they are not valuable resources. The skin becomes dry and damaged after changing color (Blokhin et al., 2022). Although the lesions are often limited to the upper portion of one limb, there have been instances in which they have spread to other parts of the body, including the chest, udder, diaphragm muscle, base of the tongue, and heart muscle (Abreu et al., 2018).

The feet, legs, and tongue are commonly the locations of predilection (Sousa et al., 2024). Lesions sometimes appear in more than one of these areas in a single animal. This illness advances quickly, and 12–36 hours after symptoms start to appear, the animal passes away silently. Many animals die without showing any symptoms (Hansford, 2020).

Sheep

Sheep with blackfoot lesions in their leg muscles have a rigid walk and are immobile because of extreme lameness in one or, more frequently, multiple legs (Disasa et al., 2020). Some animals may exhibit moderate lameness, whereas others may have severe enough lameness to impede walking. It is rare for subcutaneous edema to occur, and gas crepitation may not be noticeable before death. Skin darkening is possible although gangrene and skin necrosis are not (Useh et al., 2003). The wool from the afflicted parts of sheep can be easily removed. The muscles and other tissues are dark crimson or nearly black and appear rather dry and porous when the swelling is sliced. This is caused by the gases that the organisms release to separate the fibers. The muscular tissue and blood- stained exudate are bleeding in other areas. Dark lesions typically appear at the site of damage and organism entrance in sheep and goats (Sousa et al., 2024).

Large local lesions occur when infection results from a rupture in the epidermis, vulva, or vagina. Head lesions may be accompanied by nose bleeding and significant local swelling due to edema (Disasa et al., 2020). Ewes treated in the second group will have weakness, recumbency, or an enlarged abdomen because of edema and gas development in the fetus, which may be a source of C. chauvoei. Infected ewes at shearing will have the usual symptoms (Tagesu et al., 2019). Anorexia, despair, and high temperatures were present in every case, and death occurred quickly. Cattle and sheep with C. chauvoei-associated cardiac myositis are typically found dead (Disasa et al., 2020).

Horses

Horse clinical syndrome is not clearly described. Incoordination, stiff gait, and chest edema were noted. It is uncommon for wound infections caused by C. chauvoei to develop in local gas gangrene (Useh et al., 2003). However, in some situations, the infection may spread and affect a sizable region adjacent to the original lesion. After receiving intense antibiotic therapy, the majority of affected horses die, but in those that survive, the damaged tissue may slough off, leaving a big hollow that takes time to mend (Sousa et al., 2024).

Transmission

Although the bacterium enters the body through the mucosa of the digestive tract after ingesting tainted feed, blackleg is an infection spread by dirt (Santos et al., 2019). In addition to the spleen, liver, and digestive tract, bacteria can also be found in soil and pastures, where they may be present due to contaminated feed or the decomposition of disease- dead animal corpses (Ziech et al., 2018). Spores that become embedded in healthy tissue and proliferate due to trauma or toxemia are the precursors of true blacklegs (Echenique et al., 2018).

In sheep, the illness is nearly invariably wound infection, whereas in cattle, it typically manifests without a history of trauma (Disasa et al., 2020). Infection of skin wounds during shaving and cutting and the navel after delivery can lead to the formation of local lesions (Ziech et al., 2018). Serious outbreaks can emerge from infecting ewes’ vulva and vagina during parturition. The disease has affected flocks of ewe ewes and young rams as young as 1 year old, typically due to infection of skin wounds from fighting (Underwood et al., 2015). Sheep can experience epidemics after receiving an enterotoxemia vaccine. It is possible that the formulated vaccination induces sufficient tissue damage to allow the growth of latent spores of the organism to grow (Sousa et al., 2024). The lamb fetus exhibits a unique phenomenon. This disease is not transmitted directly from sick animals to healthy animals through contact.

Although this disease is found worldwide, it is usually limited to specific farms or pastures (Morrell et al., 2022). It is improbable that C. chauvoei grows in soil; however, due to its location, it is believed to be spread via the soil (Tagesu et al., 2019). The bacteria can be recycled through fecal contamination of the soil and are easily grown in the digestive system of cattle (Manyi- Loh et al., 2016). After exposure to the environment, C. chauvoei quickly produces spores that live for lengthy periods (years) in the soil (Daly et al., 2009). Numerous animals are susceptible to this illness, which often manifests in a matter of days. The illness is widespread in the region, particularly after floods, when areas can range in size from clusters to isolated fields formed when blackleg reaches 100% (Jamil et al., 2022).

Risk factors

Most incidences of blackleg illness in cattle occur during the warmer months of the year, and the disease typically manifests seasonally (Heckler et al., 2018). Depending on the age at which calves reach the sensitive age range, the peak incidence may occur in the spring or fall (Ayele et al., 2016). In some places, years with a lot of rainfall are associated with a higher prevalence (Jamil et al., 2022). There have been outbreaks after soil excavation, indicating that latent spores may be exposed and activated by soil disturbance (Morrell et al., 2022). The true blackleg disease is normally considered a disease of cattle and rarely sheep; however, outbreaks have been observed in deer and in one case in horses (Uzal, 2012). In cattle, the illness primarily affects young animals between the ages of 6 months and 2 years (Ayele et al., 2016). In the field, the disease appears to affect cattle that are growing quickly and are well-nourished the most (Abreu et al., 2018). Increasing sheep protein intake to improve their nutritional state makes them more vulnerable to blackleg illness (Sousa et al., 2024). There is no upper age limit for sheep.

Economic impact

In many regions of the world, blacklegs cause cattle farmers to suffer significant financial losses. Although outbreaks still occasionally occur in herds of cattle that have received all recommended vaccinations, vaccination often prevents significant outbreaks (Abreu et al., 2017). It is common for several animals to become afflicted within a few days of the sickness occurring (Nampanya et al., 2019).

Treatment

Penicillin treatment and surgical removal of the lesions, including fasciotomy, are recommended for infected animals that are not in danger of death (Sousa et al., 2024). Because lesions are so widespread, recovery rates are low. A longer acting formulation should be administered after intravenous crystalline penicillin in large dosages (44,000 IU/kg BW) (Guizelini et al., 2020). Treatment with blackleg antiserum is unlikely to be effective unless extremely high dosages are used (Tagesu et al., 2019). Because clostridial myositis progresses rapidly, treatment is rarely effective. Supportive care, vigorous surgical cleaning to allow aeration, and the use of antimicrobials (the preferred medication is procaine penicillin) around the afflicted tissue may be helpful (Li et al., 2014). In most situations, the prognosis is not good. Given the extent of the lesions, treating afflicted animals with penicillin makes sense if they are not dying, although the outcomes are typically excellent (Guizelini et al., 2020). Starting with intravenous crystalline penicillin, large doses should be administered. Longer acting formulations should then be administered, some of which should be delivered to the afflicted tissue if it is water-absorbent (Useh et al., 2006). Blackleg antiserum is unlikely to be of much use in treatment unless administered in large quantities (Disasa et al., 2020).

Vaccination

Vaccination is a crucial part of the health management of many livestock production enterprises and can prevent blackleg illness. The bacterins used in conventional blackleg vaccinations are derived from formalin-fixed cultures of C. chauvoei and are typically offered in polyvalent formulations with additional clostridial ingredients (Khiav and Zahmatkesh, 2021). Measurements of antibody titers in vaccinated animals or anecdotal evidence are the main sources of evidence supporting the effectiveness of these vaccinations. Nevertheless, very little information is available on clinical studies of this vaccination in animals. Nonetheless, scant data indicate that this vaccine is 50%–100% efficacious against experimental challenges with C. chauvoei and approximately 100% effective in preventing blackleg disease after natural exposure (Khiav and Zahmatkesh, 2021). The first vaccination against blackleg disease is typically advised at 2 months of age, with a booster shot administered 4 to 6 weeks later due to the disease’s age distribution. After that, booster shots should be given every year or every 2 years until the cow is 2 years old (Sousa et al., 2024). This toxin is a viable candidate for a novel vaccination against blackleg disease because recombinant CctA protects guinea pigs against aggressive strains of C. chauvoei (Frey et al., 2012). Neuramaminidase from C. chauvoei is another potential candidate for vaccine manufacturing (Uzal et al., 2022). It is hoped that this antigen will exert a greater protective effect against blackleg disease when combined with one or more polypeptides derived from the toxins generated by C. chauvoei. Furthermore, antibodies against C. chauvoei flagella trigger a defense immunological response (Kijima-Tanaka et al., 1994). It has been proposed that the incidence of blackleg disease can be decreased by reducing stress and trauma, particularly in younger cattle (Abreu et al., 2018).

Control

Blackleg disease’s possible occurrence is a crucial consideration for management and prevention strategies. In addition, funded vaccination programs can help minimize the occurrence of blackleg illness and consequently reduce environmental pollution caused by C. chauvoei spores, especially in high-risk locations (Salvarani and Vieira, 2024). Blackleg vaccination should be a normal operation on all properties in places where the illness is known to occur (Sousa et al., 2024). Vaccination is recommended for susceptible cattle in endemic areas. Because of the possibility of infection spreading through dead cattle that may settle after floods, animals in flooded areas or along watercourses should also receive routine vaccinations (Khiav and Zahmatkesh, 2021). Beef calves are usually vaccinated when they are ear branded, usually at one to 4 months of age.

The control of cases of blackleg disease through an official surveillance program requires coordination (Konteh et al., 2023). Providing incentives for reporting incidents can encourage farmers to report every incident. Alternative ways to prevent and control blackleg disease include specialized pasture management practices, such as artificial drainage in pastures (Ziech et al., 2018). In fact, a recent study demonstrated that cases of blackleg disease tend to be concentrated in regions with low water permeability (Santos et al., 2019). Blackleg disease can be controlled by destroying the carcass and burning the top layer of soil to eradicate any remaining spores. Many spores are dispersed by scavengers, wind, and rain (Wu et al., 2024). Relocating cattle away from contaminated areas is essential for blackleg control. Cattle on pastures with spores should be shifted (Abreu et al., 2018).

Conclusion

In conclusion, blackleg disease, primarily affecting cattle due to the anaerobic bacillus C. chauvoei, poses significant risks and financial losses to the livestock industry. Effective vaccinations have reduced its prevalence, but outbreaks can still occur, especially in inadequately vaccinated animals.

Acknowledgments

The authors thank the Faculty of Veterinary Medicine, Universitas Airlangga.

Author’s contributions

WT, ARK, BWKW, and MHE drafted the manuscript. IBM, DAAK, WT, and JYHT revised and edited the manuscript. MKJK, SW, IF, and RZA took part in preparing and critical checking this manuscript. IFM, KAF, BPP, and SR edit the references. All authors have read and approved the final manuscript.

Conflict of interest

The authors declare no conflict of interest.

Funding

The authors would like to acknowledge Ebonyi State University, Abakaliki, Nigeria, and Lembaga Penelitian dan Pengabdian Masyarakat, Universitas Airlangga, Indonesia, for their support. This study was partly supported by the International Research Consortium, Lembaga Penelitian dan Pengabdian Masyarakat, Universitas Airlangga, Surabaya, Indonesia, in the year 2024 (grant number: 171/UN3.LPPM/PT.01.03/2024.

Data availability

All references are open-access, so data can be obtained from the online web.