Chicken infectious anaemia is a disease causing great economic loss to the poultry industry by creating a health issue in commercial poultry¹,². Chicken infectious anaemia (CIA) virus, the etiological agent is a potent immunosuppressive virus, belongs to the genus Gyrovirus under family Anelloviridae³. The CIAV is a very small (23.5 to 25 nm), naked, icosahedral virus with a genome consists of a single molecule of circular singlestranded ambisense or negative sense DNA of 2.3 kb⁴. The genome encodes for 3 viral peptides of 51.6 (VP1), 24 (VP2), and 13.6 (VP3) kDa, respectively from single major transcript of 2.0 kbp size from three overlapping reading frames (ORF1, 2 and 3). The neutralizing antibodies are produced against VP1 and VP2 proteins mainly⁵,⁶. The virus was first reported and isolated from Japan following a vaccine contamination⁷. There is no antigenic difference among the strains of CIA virus, except a strain CIA virus-7 reported from the USA which could be a prototype virus of serotype 2⁸. The disease can affect all age groups but the clinical disease is mainly develops in birds below 3 weeks of age, which is characterized by generalized lymphoid atrophy, severe anaemia, reduced weight gain, immunosuppression and vaccination failure²,⁹. Due to the severe immuno-suppression, chicks become highly susceptible to secondary infections and become susceptible to various infectious and non-infectious diseases¹⁰.

In India, CIA was first reported in 1994 while investigating an outbreak of multifactorial aetiology in young growing chickens in poultry belt of Namakkal, Tamilnadu¹¹. Later the existence of CIA was confirmed from three states: Tamil Nadu, Maharashtra and U.P. by producing disease using infected materials¹². The virus detection, isolation and antibody detection in the poultry belts from different states of India further substantiated the occurrence of this disease². However, there is still lack of information regarding Chicken infectious anaemia (CIA) viruses circulating in North Eastern part of India¹³. As a result, the present study was undertaken to gather information on the prevalence, pathology, molecular diagnosis of CIA virus from commercial poultry population of Mizoram state.

A total nos. of 185 cases from 16 farms comprising tissue sample of thymus, spleen, liver, bone marrow and bursa of Fabricius were tested by PCR for detection of VP2 gene of CIAV. Out of 16 farms, 10 farms were tested positive for CIAV. A 713 bp fragment of CIAV specific VP2 gene was amplified (Fig. 1).

Affected birds showed stunted growth, ruffled feathers and were dull and depressed. Comb, wattles and visible mucous membranes appeared pale and anemic. Areas of diffuse haemorrhages were seen under the skin of wings and also in the neck and the head regions. Although, the disease was recorded in the age group between 4–12 weeks, severity of the clinical sign was observed higher in the birds of 4 weeks of age. Secondary infection was another major alteration that was observed in most of the affected farms. In three farms, CIA virus was detected with severe ascites along with the clinical signs of CIA as described. Heavy Ascaridiasis was observed in another three farms along with CIA. Curled toe paralysis was frequently observed in chicken affected by CIA.

The gross lesions included poor body condition with retarded growth, while haemorrhagic dermatitis was found to be most prominent in the wings (Fig. 2). Marked anaemia was recognized by pale musculature and viscera in de-skinned carcass. Subcutaneous haemorrhages were also observed in the lateral and medial surface of the thigh and near the atlanto-occipital joint (Fig. 3). The most consistent lesion observed was atrophy of the thymus (Fig. 4). In few cases, thymus was found to be oedematous, congested and showed pinpoint haemorrhages. The spleen was congested and atrophied in the majority of cases. On the other hand, bursa of Fabricius was oedamtous and appeared translucent on serosal surface in few cases and atrophied in majority of the cases. On dissection, bones of the affected birds appeared to be pale, white and had watery bone marrow (Fig. 5). Secondary complications such as, ascites, Ascaridiasis and catarrhal enteritis were observed in birds from five of the CIA virus affected farm.

Microscopic examination of liver depicted degenerative changes in hepatocytes, congestion with Fabricius (Fig. 7). Clear areas of inter follicular oedema with congestion were also observed. Thymus showed congestion of interlobular septae, areas of severe lymphoid atrophy resulting in loss of demarcation between cortex and medulla (Fig. 6). Intranuclear eosinophilic inclusion bodies that were considered as pathognomonic lesion from CIA has been observed in infiltrating macrophages in the depleted areas of the thymus (Fig. 8). Decalcified sections of bone revealed hypoplasia of haematopoietic cell lineage in the bone marrow. Eosinophilic intranuclear inclusion bodies were also found in the bursa of Fabricius and bone marrow.

The present report contains a description of some clinico-pathological observations on the natural outbreaks of Chicken infectious anaemia in the Northeastern region of India. The aim of this work is to confirm the outbreak of this disease which had not been reported from this region. While the poultry industry is growing at 8–10% per annum at national level, the progress of poultry industry in north eastern region is moving in slower pace due to outbreaks of different disease conditions with meagre veterinary support plays an important role to augment poultry production in the above mentioned region¹⁷. The most likely interpretation of this condion is perhaps due to outbreaks of immune-suppressive viral diseases like CIA that affects chickens of 3–4 weeks of age. This results in poor weight gain, reduction in dilated sinusoidal spaces. The spleen showed severely depleted areas of lymphoid tissue in white pulp. There were infiltrations of histiocytes in the white pulp around the sheathed artery of spleen. Severely depleted areas of lymphoid tissue were also observed in the follicles of the bursa of haematocrit values, anaemia, aplasia of the bone marrow and generalized lymphoid atrophy particularly of the thymus leading to immune-compromise and increased mortality¹⁸,¹⁹.

The causative agent CIA virus is ubiquitous and highly resistant in nature, can persist in the environment for a long time²⁰. The information on the prevalence, epidemiology, pathology and diagnosis of this disease in poultry of NER, India is yet to be uncovered. After having employed molecular technique, we confirmed the presence of CIA in this region and unequivocally, for the first time from this area and in fact it is omnipresent.

Following clinico-pathological studies, outbreaks of the disease were confirmed by the detection of VP2 gene of CIAV in tissue lesions of thymus, bone marrow and spleen. High prevalence of the disease with clinical manifestations like stunted growth, anaemia and haemorrhagic dermatitis clearly indicates a possible economic impact to the poultry farmers of Mizoram due to slow growth and delay in slaughter weight, carcass condemnation and mortality. The morbidity and mortality rate recorded in different affected farms were mainly determined by the additional complications, like ascites, ascaridiasis and enteritis. Mortality rate rose upto 60% in a CIA affected farm, when the infection was complicated by ascites and Ascaridiasis. Despite regular practice of de-worming, heavy ascaridiasis was recorded in three broiler flocks in the age group of 3–4 weeks that were affected by CIA. It was thought to be due to a possible indication of immunosuppression caused by this virus in the affected birds. Curled toe paralysis along with the paralysis of the legs was frequently observed in CIA affected farms particularly when they are of three to five weeks of age. This result is in agreement with that of earlier workers²,¹⁸,²¹,²²,²⁴.

The characteristic gross lesions including atrophy of thymus and other lymphoid tissues’ haemorrhage in muscles’ pale and watery bone marrow observed presently in the affected birds were similar to those observed by earlier workers²,²³. However, occurrence of cutaneous haemorrhages in atlanto-occipetal region of the neck region has not been reported in the past.

Direct evidence of bacterial invasion was not obtained but the presence of degenerative changes in the liver, depletion of lymphoid tissues like thymus, depletion of the haematopoietic cell lineage in bone marrow and eosinophilic intranuclear inclusion bodies in thymus, bursa of Fabricius and bone marrow were taken to be circumstantial evidence of any other unknown factor(s) to be involved like overall management, nutritional deficiency etc. So far as we are aware, this is the first occasion on which confirmed a very high prevalence of CIA in the poultry population of Mizoram, NER, India. Therefore, further studies may be initiated to study the economic impact on poultry farmers of CIA and the underlying reasons to reduce mortality by improving biosecurity, prevention of vaccination failure and secondary infections, to help farmers to enhance growth and prevent losses from carcass condemnation.

Authors are thankful to the authorities of the Central Agricultural University, Imphal, Manipur, for providing necessary support and laboratory facilities to carry out this research.

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