Paratuberculosis (Johne’s Disease) (JD)

Paratuberculosis (Johne’s Disease) or JD is mainly recognized by chronic granulomatous enteritis, emaciation and the resulting death. At very first, the disease was identified by Johnes and Frothingham in 1895.

Etiology

Paratuberculosis (Johne’s Disease) is caused by Mycobacterium avium subspecies paratuberculosis (Map). It is a non-chromogenic, facultative intracellular acid fast, non-spore forming, non–motile, curved or straight or rod shaped and aerobic organisms.

Map is usually identified by two forms; first is cell wall deficient (CWD) form and the second is cell wall competent (CWC) form. The species mycobacteria is broadly classified into slow growers and rapid growers. The slow growers are subdivided into pigmented and non pigmented group.

Structure of cell wall of Mycobacteria is about 40% of the total dry weight of the mycobacteria is made of lipid rich cell wall. It consisted of four different layers such as inner plasma membrane, immediate electron dense layer, electron transparent layer and outer membrane.

The propensity of mycobacteria to clump or aggregate in culture media is attributed to the hydrophobic nature of their cell wall and the waxy material, lipoarabinomannan (LAM) present in the cell wall is associated to the acid fast property of mycobacteria.

Epidemiology

Prevalence of infection

In Western Norway, paratuberculosis is described as endemic in goats, at the same time reported the occurrence of highest sero-prevalence in roe deer and red deer due to distribution of infection between wild and domestic animals.

The disease has been reported from various parts of the world including India.

Economic Impacts

Paratuberculosis is reported to incur substantial financial losses to livestock industry.  These losses have generally been estimated by calculating the reduction in milk production, lean meat production, culling and early replacement of infected cattle.

Predisposing factors

Pregnancy, parturition and lactation are presumptive influencing factors to enhance, the development of the disease in cattle.

Source of Infection

Map survives for prolonged periods at low temperature outside the animal host and can remain viable for several months in contaminated water and soil.

Transmission

Oral ingestion of feed, water and milk contaminated with infected feces is the principal route for transmission. A single gram of infected feces can potentially infect thousands of neonates.

There is interspecies transmission between cattle and sheep and goats. Infected cows, regardless of their disease status, excrete 5-8 cfu of organisms per 50 ml of milk.

Vertical transmission of paratuberculosis is reported to occur through reproductive tract, fetus, blood, milk and colostrums of animals in advanced stages of infection.

Eventually, horizontal transmission should also be considered important.

Nematode larvae and nymphs of oriental cockroach acts as mechanical (passive) vectors.

Host Affected

Natural infection has been reported in cattle, sheep, goats, rabbits, antelopes and bison. Para tuberculosis in water buffaloes was detected from India. Calves younger than 4 month are highly susceptible than adult cattle.

Pathogenesis

Calves acquire infection as soon as they born, both by passive and active routes. Most of them able to eliminate the infection before they become clinically symptomatic. Due to persistence of organism in their body, which would results in a chronic disease.

Persisted organism closely attached, to the glycol-proteins of the intestinal mucosa so as to escape from innte host immune mechanisms elicited by various physical, chemical and enzymatic processes.

M cells, present in dome epithelium, that lack brush border, mucosal epithelium, digestive enzymes and surface mucous provide an easy access for the close attachment of the organisms and let them to cross the intestinal epithelium, through simple transcytosis process.

Integrins (CR3), expressed on the apical surface of the M cells, acts as a bridge between fibronectin (FN) and fibronectin attachment protein (FAP) of map during phagocytosis process.

Within the intestinal mucosal epithelium the bacteria is taken up by sub-epithelial macrophages where they can multiply and destroy macrophages.

Pathogenesis and clinical signs would develop as the bacteria spread to adjacent cells. The whole process, from the time of uptake of Map by M cells and to phagocytosis by macrophages is completed in 20 hours.

Secretion of pro-inflammatory cytokines and polysaccharide complexes by the infected host cells tend to produce the granulomatous enteritis especially at the terminal part of the intestine and local lymph nodes, edema and progressive thickening of both the intestines, due to infiltration of the infected tissues with a large number of inflammatory cells.

Clinical Signs

Calves are infected as young following ingestion of milk, feed and water contaminated with M.paratuberculosis.

This causal agent is capable of living in animals for years without showing any clinical signs.

Affected cattle normally appears to be healthy and bright looking without causing any changes in their appetite , nevertheless, they exhibit some of the symptoms such as decreased in milk production, loss of body conditions, and diarrhoea in the early stage cachexia and hyporpotenemia which leads to death in the advance stage.

In small ruminants, the course of the disease may vary from rapid to prolonged periods of time and the symptoms are much more similar to cattle, excepting less frequent diarrhea in the former species.

Infected animals, during the clinical phase, pass as high as 1010 organism per gram of feces.

Depending on the extensive nature of infection, the disease has been grouped into different stages:

• Stage I – silent infection; includes both young and adult with no shedding of bacteria.
• Stage II – subclinical excretory phase; adult carriers without clinical signs.
• Stage III – clinical excretory phase; animals with characteristic signs
• Stage IV – advanced clinical disease.

Necropsy Findings

Microscopic lesions of JD is predominantly divided into two; Lepromatous and tuberculoid granulomatous lesion.

Lesions are classified into four types based on the pathological features manifested by the Map infected goats.

Diagnosis

• Based on clinical signs and necropsy findings.
• Detection of acid fast bacilli from dung sample, intestinal mucosa and associated lymphnodes in infected animals are the most essential confirmatory diagnostic method of Para tuberculosis.
• Isolation of organism by culture. Solid media which includes, the egg based Lowenstein- Jensen medium(LJ) and Herold’s egg yolk medium (HEYM).
• The culture method was described as ‘gold standard’. culture is time consuming, may require up to 3 months for incubation and labor intensive.
• Fecal culture test is most sensitive and could be used in herd level testing. In BACTEC radiometric culture, the growth and recovery of colonies is observed to be greater, more sensitive and time saving than the conventional culture system .
• Cellular immunity is the first and strongest host response elicited during the early stages of mycobacterial diseases, is essential for the control of infection.
• Similarly in Para tuberculosis infected animals develops a strong CMI response in response to paucibacillary form (tuberculoid type) and a weak CMI to multibacillary form (lepromatous type).
• This cellular response is usually characterized by delayed type IV hypersensitivity (DTH) / intradermal test, lymphocyte transformation assay, migration inhibition test (MIT), (IFN-Y assay), and interleukin 2-R assay.
•  Invivo–intradermal test is a very old technique used to measure sensitized T-lymphocytes response to an antigen.
• It is a field test using 0.1-0.2ml of johnin PPD at caudal fold or the neck .
• The reactions of the animals are measured from a difference between the initial and the final thickness of the skin.
• Animal showing skin thickness of above 4mm after 72 hours of injection is considered positive.
• An alternative method of measuring CMI response is the gamma interferon assay, a invtro test, is a sensitive test and it measures the release of interferon by sensitised lymphocytes and is considered effective in screening and culling of infected herd.
• CMI response can also be measured by the expression of interleukin 2 receptor (IL-2R) on lymphoblasts by flow cytometry analysis.
• There are several methods like Complement Fixation (CFT) Test, Agar Gel Immunodiffusion (AGID) Test and Enzyme Linked Immunosorbent Assay (ELISA), used in the assessment of humoral immune response of paratuberculosis.
•  In which CFT test has been considered a best screening test for cattle meant for international export.
• Molecular diagnosis by DNA probes may be of use in identifying clinically important bacteria.
• But for the bacteria like M.paratuberculosis, , the use of species specific DNA probe may be essential.
• To date only IS900, F57 and HspX genes that are specific to this disease have been identified from its genome.

Sample collection

Dung sample, intestinal mucosa and associated lymphnodes in infected animals are the most essential confirmatory diagnostic method of Para tuberculosis.

Differential Diagnosis

• Tuberculosis
• Parasitic disease
• Salmonellosis
• Colibacillosis Rota viral enteritis

Treatment

There is no treatment and antibiotics are available to control paratuberculosis. Temporarily diarrhoea can be controlled by some antibiotics. Combined use of Streptomycin, Isoniazid and Rifambicin is suggested in some countries.

Prevention

There are two vaccines commercially available used in cattle , sheep and goats against Para tuberculosis in New Zealand; a live attenuated freeze dried vaccine which was Introduced in New Zealand and the killed vaccine has been involved in the practice .

The killed vaccine is  found to reduce number of organism excreted, spread of infection, and development of lesion in the body.

In Para tuberculosis, the development of immune mechanism is bipolar in nature. A strong cellular immune response (CD4+Th1 cells) develops during the earlier period of infection, is the prime response by which the progression of infection is controlled.

CMI wanes and simultaneously provoke the humoral response ( CD8+ Th2 cells)at the later stage.

As the immune system in calves is not completely developed they are highly susceptible to disease as compared to adult cattle.

Control

The control of paratuberculosis mainly depends on the early diagnosis of disease by sero-surveillance, quarantine of suspected and culling of infected animals, eradication by vaccination and maintenance of hygiene in the farm premises.

Generally surveillance relies on serological tests such as CFT and ELISA. To date, as antibiotics are proved to be ineffective, expensive and time consuming calf hood vaccination is considered an other method to curtail the progression of paratuberuclosis from subclinical to clinical stage.

However, the long incubation period, resistant nature of the organisms to the external and internal environmental conditions , shedding through feces makes the control of paratuberculosis become difficult.

As young calves are reported to be highly at risk group of age, separation of calves as soon as they were born is recommended in the first place, in order to avoid them from getting infection from dam is the pasteurization of milk.