The Horse with Recurrent Fevers and Infections: Approach to Common Variable Immunodeficiency in the Horse

Introduction

Common variable immunodeficiency (CVID) in horses shares many of the same features as the human CVID. The aim of this review is to inform equine practitioners of the clinical manifestations and diagnostic approach for CVID in the horse in order to support timely diagnosis. We provide information on the current state of knowledge of mechanisms of CVID in the horse and highlight management options currently available. By increasing the awareness of CVID in horses, we aim to decrease the diagnostic delay and improve the welfare of equine patients with this devastating disease.

In humans, CVID is the most common form of primary antibody deficiency, although it is still considered a rare condition with an estimated incidence of 1:25,000.¹ It encompasses a heterogeneous group of immune defects characterized by hypogammaglobulinemia and failure of specific antibody production resulting in severe bacterial infections, primarily, but not limited to respiratory tract infections.¹ Complications include damage to organ systems as a result of severe or recurrent infections.¹ Additionally, autoimmune, inflammatory and lymphoproliferative conditions have a higher prevalence in CVID patients compared to the general population.² As a rare disease, there is limited knowledge of the clinical, diagnostic and therapeutic approach to CVID leading to misdiagnosis and undertreatment in human patients. In fact, CVID patients face a median diagnostic delay of 4 years.¹

As we will discuss, CVID in horses shares many of the same features as the human disease. The aim of this review is to inform equine practitioners of the clinical manifestations and diagnostic approach for CVID in the horse to support timely diagnosis. We provide information on the current state of knowledge of mechanisms of CVID in the horse and highlight management options currently available. By increasing the awareness of CVID in horses, we aim to decrease the diagnostic delay and improve the welfare of equine patients with this devastating disease.

Background

Common variable immunodeficiency (CVID) in the horse is characterized by late-onset B cell depletion and/or dysfunction resulting in hypogammaglobulinemia and recurrent bacterial infections including respiratory diseases, cholangiohepatitis, septicemia and/or meningitis.^3,4 It is a rare, although likely underrecognized condition in horses, with more than 100 confirmed cases identified to date by the Equine Immunology Laboratory, Cornell University.⁵

The disease was first reported in a 12-year-old Quarter Horse mare presented for lethargy, anorexia, and intermittent chronic nasal discharge of three weeks duration.⁴ The initial case report of CVID highlights the salient historical, clinical examination, and laboratory findings in horses with CVID, which are discussed here to illustrate the systematic approach for diagnosing CVID. The adult mare was noted to be mildly febrile (rectal temperature 101.1F), with a neutrophilic leukocytosis and left shift. Serum biochemical abnormalities included elevated alkaline phosphatase, aspartate aminotransferase and gamma glutamyltransferase with concurrent hypoglobulinemia. She was diagnosed with Streptococcus equi subspecies zooepidemicus guttural pouch empyema that responded poorly to medical and surgical management. She subsequently developed a surgical site infection, severe diffuse cellulitis and abscessation of the subcutaneous tissue of the ventral cervical region that cultured positive for Streptococcus equi subspecies zooepidemicus, and bilateral maxillary sinusitis. The owner declined liver biopsy, but a presumptive diagnosis of bacterial cholangiohepatitis secondary to dissemination of the streptococcal organism was made.

Immunological testing revealed trace serum IgG and IgM concentrations and markedly low B lymphocyte distribution in peripheral blood using flow cytometric analysis. Furthermore, serum samples obtained before and 20 days post-vaccination with tetanus toxoid were analyzed for tetanus antibody titers as a marker of antibody production. The results revealed no specific antibody production, confirming the impaired humoral function.

At the owner’s request, the mare was discharged from the hospital six weeks after admission for continued treatment with antimicrobials (SMZ-TMP and rifampin) and antiinflammatories (phenylbutazone). She was readmitted 4 weeks after discharge due to sudden deterioration in condition. She was noted to be weak, hypothermic and have an abnormal mentation. She had developed a severe neutrophilic leukocytosis with markedly elevated liver enzymes. Differential diagnosis included septicemia and hepatic encephalopathy. She was euthanized and a full necropsy was performed. Major findings included mild, focal, unilateral guttural pouch empyema, mucopurulent and suppurative bronchitis, and lymphoplasmacytic hepatitis and cirrhosis. There was also generalized lymphadenopathy and loss of normal lymph node histological architecture characterized by absence of lymphoid follicles.

Immunohistochemistry performed with B cell monoclonal antibodies revealed no positively stained cells in the bone marrow and spleen, and only occasional positive stained cells in the lymph nodes.⁴

Epidemiology

Since the initial CVID case report was published in 2002, additional case reports and case series have been published that underscore the heterogeneity of the demographics, phenotypic features and pathogens associated with CVID in horses.^3,6,7 The disease distribution suggests a non-familial pattern without obvious shared environmental factors. The prevalence of CVID in horses is unknown, however, it represents 13.1% of the accessions to the Equine Immunology Laboratory that offers testing specific for humoral immunodeficiencies. The median age at diagnosis was 12 years of age (range 2–27 years).⁵ In most reported cases, the clinical signs began one to two years before hypogammaglobulinemia was measured, indicating a delay in the diagnosis of CVID. Affected patients were unrelated horses of both sexes with 57% male and 44% female. Breeds affected included Thoroughbred and Thoroughbred crosses, Quarter Horse and Quarter Horses crosses, Warmblood and Warmblood crosses, Arabian, Paint, Pony, Friesian, Irish Draft, Paso Fino and Argentine Polo. Cases were located throughout the United States, with 1 case in Australia, and only single cases per herd have been diagnosed.⁵

Approach to diagnosis

In reviewing the commonalities among CVID patients, frequent or sporadically recurring fevers were found in almost all cases. When approaching cases with this history, the equine practitioner should have an increased index of suspicion of an underlying immunodeficiency. The most common clinical presentations of horses with CVID are recurrent respiratory infections, meningitis and/or ataxia, cholangiohepatitis, infectious colitis, infectious dermatitis and/or severe gastrointestinal (GI) parasitism.⁵ Other clinical manifestations include weight loss, muscle mass loss or poor body condition. Meningitis and multiple organ system involvement (e.g. pneumonia and bacterial cholangiohepatitis) may result from septicemia. Meningitis is an uncommon finding in adult horses and should raise the index of suspicion of CVID. Meningitis presents clinically as marked depression, hyporexia, ataxia, neck stiffness and low head positioning. Cerebrospinal fluid analysis in CVID cases with meningitis has revealed a neutrophilic pleocytosis with increased protein concentrations consistent with infection.⁶

Complete blood count and serum chemistry panel abnormalities that should alert the clinician to the possibility of CVID include a persistently low globulin concentration in the presence of severe bacterial infection; a transient or persistent lymphopenia (<1,200 cells/uL) is also common but not always present (Figure 1). Some cases have elevated liver enzymes consistent with cholangiohepatitis, likely due to increased susceptibility to ascending infection from GI tract and/or hematogenous spread of infection.⁴ The findings of elevated liver enzymes combined with fever, neutrophilia, and hypoglobulinemia warrants further investigation for CVID. Depending on the duration of infection, markers of acute (increased serum amyloid A) or chronic inflammation (mild anemia and hypoalbuminemia) may be present.⁵

Figure 1:

Diagnostic approach to CVID in the adult horse.

The diagnosis of CVID is made based on the presence of hypogammaglobulinemia defined as IgG < 1,000 mg/dL in adult horses along with supporting clinical signs of infections (Figure 1). Serum IgM concentrations are variable in horses with CVID. Commercial tests for serum IgA in horses are not currently available, but previously, horses with CVID could have normal serum IgA concentrations that decreased over the course of disease.³ Serum IgE concentrations were measured in several horses with CVID and found to be undetectable or markedly low.⁵

Response to vaccination has been used as a proxy for humoral competency. Horses with CVID have no to minimal rise in tetanus specific antibodies 15–21 days post vaccination.^3,4 Although informative about the extent of humoral function, vaccination response tests should be interpreted cautiously because of individual variations in humoral response to different types of vaccines.⁵ Hence, reliance on serial measurement of serum IgG and IgM concentrations provides the best assessment of disease progression and extent of humoral immunodeficiency in affected horses.

Approach to rule out other causes of recurrent infections

For the equine practitioner faced with a patient with recurrent infections, important considerations include the patient’s signalment and clinical presentation that can help rule out other causes of recurrent infections. Causes other than CVID that may lead to recurrent infections include infections with highly pathogenic and/or antimicrobial resistant organisms. Culture and sensitivity of the appropriate tissue/fluid samples are useful in determining the pathogen(s) responsible for the infection.

Horses with malnutrition, overcrowding, parasitism, and poor management may be prone to recurrent infections in the absence of a primary immunodeficiency. The practitioner is advised to investigate the herd’s environment when faced with a horse with recurrent infection.

Epithelial damage due to viral infections, catheter placements, wounds, surgical site infections and history of severe GI disease causing disruption of the GI mucosal barrier are additional risk factors for recurrent fevers and infection in the absence of a primary immunodeficiency.^8,9 Comprehensive clinical assessment will aid in ruling out these causes of infection. Serial measurements of hypogammaglobulinemia are recommended to confirm the diagnosis of CVID in horses with subacute GI disease and GI mucosal damage where immunoglobulin loss (not impaired production) may be the cause of mild, transitory hypogammaglobulinemia. Accompanying clinical findings may include mild colic, diarrhea, hypoalbuminemia and neutropenia. A diagnosis of CVID is supported when the hypogammaglobulinemia does not resolve with resolution of the GI disease.

Another risk factor for recurrent infection is treatment with immunosuppressive therapies. Previous or current treatment with glucocorticoids and other immunosuppressive medications should be documented. While treatment with immunosuppressive medications does not rule out concurrent CVID, it can be a risk factor for a patient developing recurrent infections even in the absence of a primary immunodeficiency. In some of the reported cases of CVID, treatment with glucocorticoids was noted, increasing the clinical susceptibility to a wider range of pathogens and complicating diagnosis.⁵ Obtaining a complete and detailed medical history is recommended to help construct a disease timeline in complex cases.

In foals and young horses, it is important to rule out transient hypoglobulinemia of the young. It may present similarly to CVID, but generally occurs in horses that are less than two years of age. In this condition, foals and young horses have a normal distribution of circulating B cells, but normal antibody production levels are not achieved until 2–3 years of age. Serial measurement of IgG and IgM concentrations several times per year is recommended to assess whether humoral competency is achieved. Continued hypogammaglobulinemia without improvement supports the diagnosis of CVID.⁵

At the other end of the age spectrum, horses with pituitary pars intermedia dysfunction (PPID) are prone to recurrent infection due to immune dysregulation.¹⁰ In older horses (>15 years) with recurrent infection, ruling out PPID with baseline adrenocorticotropic hormone (ACTH) or thyrotropin-releasing hormone (TRH) stimulation test is recommended.¹¹ PPID can co-occur with CVID, and in horses with PPID that have characteristic clinical features (fevers and recurrent infections) and laboratory findings (low-normal to hypoglobulinemia) of CVID, immunological testing is recommended including serum IgG, IgM and peripheral blood lymphocyte immunophenotyping to rule in or out a primary immunodeficiency.⁵

Additional testing - peripheral blood lymphocyte immunophenotyping

Lymphocytes circulate through blood, lymph and lymphoid tissue, therefore, peripheral blood lymphocyte immunophenotyping can be used as an indicator of immune activity in lymphoid tissues (Figure 1). In 100 horses diagnosed with CVID, 63% had a persistently low B cell distribution less than 6% of the total circulating lymphocytes, indicating B cell deficiency or depletion. Normal reference intervals (median and range) vary among laboratories but, in general, for CD19 B cells are 10.2% (7.7–12.7%) and for CD21 B cells 10.7% (6.5–15.9%). In the remaining 37% of horses with CVID, the B cell distribution was within or markedly above the reference range, indicating B cell presence, but suggesting altered B cell function.⁵

Most of the horses diagnosed with CVID had a relative increase in the distribution of circulating CD8 T cells and/or CD4 T cells. T lymphocyte function and ability to stimulate B cells appeared intact in a cohort of horses with CVID based on in vitro testing and expression of interferon-gamma and CD40-ligand in stimulated cells compared to healthy horses.³

Approximately 5–10% of horses with CVID present with mild to moderate CD4 T cell lymphopenia below 50% of lymphocytes (reference interval 59.7–69.1%), leading to a CD4:CD8 T cell ratio <2.5 (reference interval 2.5–4.5) (Figure 1). In these cases, infection with fungal agents or intracellular bacteria is common. Similar findings may be observed when glucocorticoid administration is part of the clinical history.³

In human patients with CVID, those with CD4 T cell lymphopenia are more likely to present with autoimmune, immune-mediated and lymphoproliferative disease.^12–14 This association has not been found in horses with CVID. However, a subset of horses with CVID do have concurrent disorders of immune dysregulation as discussed below.

Additional testing - Post-mortem findings and histopathology

The post-mortem findings help to define the lymph node histopathological findings in CVID cases with paucity or presence of B cells (Figure 1). The post-mortem findings were available for a subset of the 100 horses diagnosed with CVID. Lesions associated with the infections were identified and corroborated the clinical diagnosis. In addition, lesions associated with vasculitis, uveitis and lymphoma were identified in a small number of cases with concurrent autoimmune, immune-mediated or lymphoproliferative diseases.

Cases with low circulating B cells commonly showed abnormal paracortical and cortical distributions of lymphocytes in lymph nodes, with absence of primary and secondary B cell follicles, and occasional B cell clusters. Lymph nodes ranged in size from small to enlarged based on the presence or absence of infection and/or T cell lymphoproliferative conditions. Immunohistochemistry also revealed B cells were absent in bone marrow and spleen and absence of plasma cells in lymphoid and mucosal tissues. Cases with circulating B cells had present germinal centers or clusters of B cells and plasma cells in lymphoid and mucosal tissues.

Equine practitioners should be advised that finding normal lymph node histology and the presence of B cells does not rule out CVID in horses with hypogammaglobulinemia and supporting clinical history.⁵

Complications and concerns - opportunistic infections

Infections with opportunistic organisms are found most commonly in horses with CVID (Figure 1). Encapsulated bacteria are often identified in these patients because they require opsonization with both antibodies and complement for effective phagocytosis and killing.⁵ For example, reported cases have had Streptococcus equi subspecies zooepidemicus respiratory infections, Staphylococcus spp., Klebsiella spp., and Borrelia burgdorferi meningitis; Salmonella spp. groups B or C and Clostridium difficile colitis; Staphylococcus spp. dermatitis; and Actinobacillus spp. peritonitis.^3,4,6,7,15

The equine practitioner is advised to submit samples for bacterial culture and sensitivity and/or PCR in suspect and confirmed CVID cases as the type of bacteria can help to determine the presence, severity and characterization of a primary or underlying immunodeficiency, affecting one or more areas of the immune system. Other opportunistic infections found in CVID cases include GI parasitism and dermatophytosis. As mentioned above, fungal (Bipolaris spp., Pneumocystis jirovecii, Aspergillus fumigatus) and intracellular bacterial (Rhodococcus equi) infections suggest that the patient may have a concomitant cellular immunodeficiency, often characterized by CD4 T cell lymphopenia.

Complications and concerns - immune dysregulation accompanying CVID

Immune-mediated diseases in human patients with CVID include autoimmune cytopenias, rheumatological conditions and vasculitis associated with inflammatory cellular immune responses and autoantibodies despite hypogammaglobulinemia.^2,16 Autoimmune and immune-mediated disorders were reported in at least 10% of the horses with CVID. Included among these conditions were vasculitis/cellulitis/lymphangitis characterized by distal limb edema, swelling, pain and/or dermatitis (Figure 1). Uveitis, either autoimmune or associated with an infectious organism were reported in several cases. Inflammatory bowel disease and recurrent urticaria were also reported.⁵ Horses with CVID and immune-mediated or autoimmune conditions require careful management as discussed below.

As in humans, humoral deficiencies and lymphoproliferative diseases can co-occur in horses. Cutaneous and nasopharyngeal forms of lymphoma, some of them characterized as T cell-rich large B cell lymphoma were described before or concomitant with hypogammaglobulinemia and the diagnosis of CVID. Hypogammaglobulinemia with or without lymphoma characterizes a humoral deficiency.⁵

Proposed mechanism of disease – Undergoing research

The mechanism of CVID in horses has been studied extensively and is partially characterized. B cells must be continuously produced in the bone marrow throughout life, starting from hematopoietic stem cells, and further differentiated in the peripheral lymphoid tissues after interaction with antigens and other cells of the immune system. In most horses with CVID, the late-onset B cell development failure appears to start in the bone marrow, leading to an obvious B cell paucity, whereas the presence of circulating and tissue B cells suggest their development but perhaps intrinsic or extrinsic faulty differentiation.¹⁷ To date, no genetic mutations have been identified and the large number of breeds of horses affected argue against an inheritable disease. Only single affected horses within a herd have been diagnosed with CVID and environmental risk factors have not yet been identified. However, recent evidence suggests that epigenetic mechanisms may be involved in CVID in horses.¹⁷

Epigenetic alterations are often associated with environmental factors that alter gene expression without changing the DNA sequence, such as DNA methylation status.¹⁸ In the bone marrow of horses with CVID and B cell lymphopenia, the expression of key early B cell signature genes (E2A and PAX5) is reduced compared to healthy horses, and this gene silencing is associated with hypermethylation of the regulatory enhancer and promoter regions of these genes.¹⁷ Thus, CVID in horses with B cell paucity appears to be associated with impaired B cell development in the bone marrow at the transition between pre-pro-B cells and pro-B cells, but the exact mechanisms triggering this aberration is still not understood.

Discussion

None of the horses with CVID recovered to normal antibody production. Short- and longterm survival is dependent on disease severity and quality of life. Horses with persistent lymphopenia, concomitant CD4 T cell lymphopenia, hepatopathy, meningitis and/or immunemediated conditions required more intensive treatment and had a reduced quality of life. Rare cases have been managed for a few to many years when serum IgG concentrations remained above 600 mg/L.⁵

Monthly immunoglobulin replacement therapy is the mainstay of treatment for humans with CVID. In equine patients, plasma transfusion is the only available source of equine immunoglobulins. While plasma transfusion may be useful in acute, severe cases, it is rarely utilized as a long-term therapy due to cost, volume needed to significantly improve serum IgG concentrations in adult horses, and short duration of providing protective antibody levels.³

Allogeneic, major histocompatibility hematopoietic stem cell transplantation has been attempted in a horse with CVID but resulted in only short-term replacement of a small population of B cells. The complexity and cost of this therapy limits its clinical applicability at this time.⁵

Routine vaccination with inactivated vaccines is recommended in horses with CVID to stimulate cellular immunity and potential remaining memory B cells. The safety of modified-live or live vaccines has not been evaluated in horses with CVID. As mentioned above, GI parasitism should be considered in horses with CVID and repeated fecal egg counts and routine administration of anthelmintics should be considered in these patients.

Horses with CVID need to be evaluated frequently for the development of infections with physical examinations and blood work. Repeated serum IgG and IgM measurements can help inform the clinician whether antibiotic therapy may be indicated. Special attention should be placed on evaluation of the respiratory tract (re-breathing examination, thoracic ultrasound evaluation, airway endoscopy, and sample collection for infectious agent testing via PCR and culture and sensitivity as indicated) and development of septicemia. Some equine CVID patients have been managed with intermittent oral antibiotics for months to years, although the potential risks of antimicrobial-induced diarrhea and bacterial resistance must be carefully considered. Severe infections such as sepsis and meningitis require hospitalization, intravenous antibiotic therapy and supportive care. In horses with CVID and concurrent CD4 T cell lymphopenia, the addition of antifungal medications may be needed.⁵

Management of immune-mediated, autoimmune and lymphoproliferative diseases in equine CVID patients present additional challenges. Medications such as glucocorticoids induce cellular immunosuppression, including CD4 T cell lymphopenia, which increases the risk of intracellular bacterial and fungal infections in horses with CVID. If glucocorticoids are required to manage co-existing conditions, the patient should be monitored closely, and administration of antibiotics and antifungal medications should be considered.

Similarly, PPID is associated with risk of immunosuppression and opportunistic infection, suggesting that horses with co-occurrence of PPID and CVID should be carefully managed with biosecurity, vaccination and other strategies to reduce infection.^10,11 Although one study found that treatment with pergolide did not reduce in vitro markers of immunosuppression in horses with PPID,¹⁰ there may be positive in vivo effects of treatment with pergolide and it is, therefore, recommended.

Summary

Equine CVID, like the disease in humans, represents a heterogeneous class of humoral immunodeficiencies. Horses presenting for recurring infections and fevers warrant further investigation into potential underlying immunodeficiencies. In horses, early recognition of the disease by the equine practitioner is paramount to managing the quality of life for these patients, for whom the long-term prognosis remains poor. Serum IgG and IgM concentrations, and peripheral blood lymphocyte immunophenotyping are the current diagnostic tests available to characterize CVID in horses. Future work to categorize horses with CVID based on their clinical, immunological, and infectious agent susceptibilities will help to further elucidate the mechanisms of disease and, hopefully, establish precision medicine options for equine CVID patients.

Clinical care points.

• Common variable immunodeficiency (CVID) in horses is a rare, non-familial condition characterized by late-onset B cell depletion and/or dysfunction resulting in inadequate antibody production and predisposition to recurrent infections.

• Diagnosis is based on supporting clinical signs, and hypoglobulinemia due to hypogammaglbulinemia (serum IgG < 1,000 mg/dL). Peripheral blood lymphocyte immunophenotyping further characterizes the disease into B cell paucity or B cell presence. Concurrent CD4 T cell lymphopenia, identified in some cases of CVID, increases risk of fungal and intracellular bacterial infection.

• Opportunistic infections are common in horses with CVID. Testing for bacterial, fungal and viral agents is recommended to aid in CVID characterization, and to guide antimicrobial therapy.

• Specific treatment is not currently available and long-term prognosis is poor, however, some cases have been managed with intermittent antibiotic therapy for months to years.

• CVID-affected horses should be routinely vaccinated with inactive vaccines to stimulate cellular immunity and memory B cells. Internal parasites should be treated with appropriate anthelmintics.

• Autoimmune, immune-mediated and lymphoproliferative conditions can occur concurrently with CVID and require careful management. Administration of glucocorticoids for these conditions can further exacerbate the risk of infection and co-administration of antibiotics and antifungal medications should be considered.

• Diagnosing and managing concurrent pituitary pars intermedia dysfunction (PPID) is recommended, as PPID presents another risk factor for opportunistic infection.

Synopsis:

Equine common variable immunodeficiency is a rare, late-onset, non-familial humoral deficiency characterized by B cell depletion and/or dysfunction resulting in inadequate antibody production and predisposition to recurrent infections. Serum immunoglobulin concentration and peripheral blood lymphocyte immunophenotyping are required to diagnose and characterize CVID in horses. Early recognition of the disease by the equine practitioner is paramount to managing the quality of life for these patients, for whom specific treatment is not yet available and long-term prognosis remains poor. An approach to the diagnosis, identification of complicating factors and management of horses with CVID is discussed.

Key points:

• Recurrent fevers and infections with hypoglobulinemia should raise the equine practitioner’s index suspicion for an immunodeficiency disorder.

• Common variable immunodeficiency (CVID) in horses is a rare condition characterized by late-onset B cell depletion and/or dysfunction resulting in inadequate antibody production (serum IgG<1,000 mg/dL) and predisposition to recurrent infections.

• CVID in horses is caused by impaired B cell development and/or differentiation, and the mechanisms involved are not completely understood.

• Specific treatment is not currently available and long-term prognosis is poor, however, some cases are managed with intermittent antibiotic therapy for months to years.

• CVID-affected horses should be routinely dewormed and vaccinated with inactive vaccines to stimulate cellular immunity and remaining memory B cells.